Ferrite nanoparticles

ABSTRACT

Methods of forming spinel ferrite nanoparticles containing a chromium-substituted copper ferrite as well as properties (e.g. particle size, crystallite size, pore size, surface area) of these spinel ferrite nanoparticles are described. Methods of preventing or reducing microbe growth on a surface by applying these spinel ferrite nanoparticles onto the surface in the form of a suspension or an antimicrobial product are also described.

CROSS-REFERENCE TO RELATED APPLICATIONS

The present application is a Divisional of U.S. application Ser. No.16/250,626, now allowed, having a filing date of Jan. 17, 2019.

STATEMENT REGARDING PRIOR DISCLOSURE BY THE INVENTORS

Aspects of this technology are described in an article “Synthesis andCharacterization of Antibacterial Activity of SpinelChromium-Substituted Copper Ferrite Nanoparticles for BiomedicalApplication” published in Journal of Inorganic and OrganometallicPolymers and Materials, 2018, Volume 28, Issue 6, pp 2316-2327, on Jun.6, 2018, which is incorporated herein by reference in its entirety.

BACKGROUND OF THE INVENTION Technical Field

The present disclosure relates to spinel ferrite nanoparticlescomprising a chromium-substituted copper ferrite, a method of preparingthe spinel ferrite nanoparticles, and methods of utilizing these spinelferrite nanoparticles for killing or inhibiting growth of microorganismsand sterilizing surfaces contaminated with biologically active pathogensand/or microorganisms.

Description of the Related Art

The “background” description provided herein is for the purpose ofgenerally presenting the context of the disclosure. Work of thepresently named inventors, to the extent it is described in thisbackground section, as well as aspects of the description which may nototherwise qualify as prior art at the time of filing, are neitherexpressly or impliedly admitted as prior art against the presentinvention.

Nanotechnology research came to prominence in the 21^(st) century as itbecame increasingly important for a broad range of practicalapplications. Ferrite nanoparticles have attracted much attentionbecause of their superparamagnetic properties as well as large surfacearea to volume ratio, which are different from their bulk counterparts[Naseri M G, Saion E B, Ahangar H A, Shaari A H, Hashim M. Simplesynthesis and characterization of cobalt ferrite nanoparticles by athermal treatment method. Journal of Nanomaterials. 2010, 1; 2010:75;and Kefeni K K, Mamba B B, Msagati T A. Application of spinel ferritenanoparticles in water and wastewater treatment: a review. Separationand Purification Technology. 2017, 188:399-422, each incorporated hereinby reference in their entirety]. Ferrites are classified as spinel,garnet, hexaferrite or orthoferrite according to their crystalstructures and magnetic properties [Singh R, Thirupathi G.Manganese-Zinc Spinel Ferrite Nanoparticles and Ferrofluids. In MagneticSpinels—Synthesis, Properties and Applications 2017. Chapter 7, InTech.http://dx.doi.org/10.5772/66522, pages 140-159]. Recently, specialattention has been given to transition metal ferrite nanoparticles withspinel structures because of their renowned magnetic, catalytic,optical, and electronic properties and high adsorption capability [SanpoN, Wen C, Berndt C C, Wang J. Antibacterial properties of spinel ferritenanoparticles. Microbial pathogens and strategies for combating them:science, technology and education. Spain: Formatex Research Centre.2013:239-50, incorporated herein by reference in its entirety]. Inaddition, because of their high permeability and good saturationmagnetization, these transition metal ferrite nanoparticles aremagnetically soft and readily magnetized and demagnetized [Mathew T,Malwadkar S, Shivanand, Pai, Sharanappa N, Sebastian C P. Oxidativedehydrogenation of ethylbenzene over Cu_(1-x)Co_(x)Fe₂O₄ catalystsystem: Influence of acid-base property. Catalysis Letters. 2003;91:217-24, incorporated herein by reference in its entirety].

Properties of ferrite nanoparticles may be tuned by modifying theirparticle size, shape, and the amount of substituted transition metalions. Spinel ferrite nanoparticles might be used in biomedicine forcancer treatment [Peng Y, Wang Z, Liu W, Zhang H, Zuo W, Tang H, Chen F,Wang B. Size- and shape-dependent peroxidase-like catalytic activity ofMnFe₂O₄ Nanoparticles and their applications in highly efficientcolorimetric detection of target cancer cells. Dalton Transactions.2015; 44(28):12871-7, incorporated herein by reference in its entirety],dopamine investigation [Reddy S. Swamy B K, Chandra U, Mahathesha K R,Sathisha T V, Jayadevappa H. Synthesis of MgFe₂O₄ nanoparticles andMgFe₂O₄ nanoparticles/CPE for electrochemical investigation of dopamine.Analytical Methods. 2011, 3(12):2792-6, incorporated herein by referencein its entirety], magnetic hyperthermia for diagnosis and treatment ofcancer, drug delivery, cellular signaling, and magnetic resonanceimaging [Céspedes E, Byrne J M, Farrow N, Moise S, Coker V S, Bencsik M,Lloyd J R, Telling N D. Bacterially synthesized ferrite nanoparticlesfor magnetic hyperthermia applications. Nanoscale. 2014, 6(21):12958-70;and Choi H, Lee S. Kouh T, Kim S J, Kim C S, Hahn E. Synthesis andcharacterization of Co—Zn ferrite nanoparticles for application tomagnetic hyperthermia. Journal of the Korean Physical Society. 2017,70(1):89-92, each incorporated herein by reference in their entirety].They may also be used for magnetic recording [Shu C, Qiao H. TuningMagnetic Properties of Magnetic Recording Media Cobalt FerriteNano-Particles by Co-Precipitation Method. In Photonics andOptoelectronics, 2009. SOPO 2009. Symposium on 2009, pp. 1-4. IEEE,incorporated herein by reference in its entirety], catalysis [Zhang L,Wu Y. Sol-Gel synthesized magnetic MnFe₂O₄ spinel ferrite nanoparticlesas novel catalyst for oxidative degradation of methyl orange. Journal ofNanomaterials. 2013, 1; 2013:2; and Waag F, Gökce B, Kalapu C, Bendt G,Salamon S, Landers J, Hagemann U, Heidelmann M, Schulz S, Wende H,Hartmann N. Adjusting the catalytic properties of cobalt ferritenanoparticles by pulsed laser fragmentation in water with defined energydose. Scientific Reports. 2017, 7(1):13161, each incorporated herein byreference in their entirety], sensing [Joshi S, Kamble V B, Kumar M,Umarji A M, Srivastava G. Nickel substitution induced effects on gassensing properties of cobalt ferrite nanoparticles. Journal of Alloysand Compounds. 2016, 654:460-6; and Zafar Q, Azmer M I, Al-Sehemi A G,Al-Assiri M S. Kalam A, Sulaiman K. Evaluation of humidity sensingproperties of TMBHPET thin film embedded with spinel cobalt ferritenanoparticles. Journal of Nanoparticle Research. 2016, 18(7):186, eachincorporated herein by reference in their entirety], water andwastewater treatment [Kefeni K K, Mamba B B, Msagati T A. Application ofspinel ferrite nanoparticles in water and wastewater treatment: areview. Separation and Purification Technology. 2017, 188:399-422],magneto-optical devices, heat absorbers and generators, energy storage,electromagnetic interference shielding, and microwave devices [Singh R,Thirupathi G. Manganese-Zinc Spinel Ferrite Nanoparticles andFerrofluids. In Magnetic Spinels-Synthesis, Properties and Applications2017. Chapter 7, InTech. http://dx.doi.org/10.5772/66522 pages 140-159;and Kefeni K K, Msagati T A, Mamba B B. Ferrite nanoparticles:synthesis, characterisation and applications in electronic device.Materials Science and Engineering: B. 2017, 215:37-55, each incorporatedherein by reference in their entirety]. Spinel ferrite nanoparticles canbe synthesized by methods such as auto combustion [Velhal N B, Patil ND, Shelke A R, Deshpande N G, Puri V R. Structural, dielectric andmagnetic properties of nickel substituted cobalt ferrite nanoparticles:Effect of nickel concentration. AIP Advances. 2015, 5(9):097166,incorporated herein by reference in its entirety], polymeric precursormethods [Gharagozlou M. Synthesis, characterization and influence ofcalcination temperature on magnetic properties of nanocrystalline spinelCo-ferrite prepared by polymeric precursor method,” Journal of Alloysand Compounds. 2009, 486: 660-665, incorporated herein by reference inits entirety], sonochemical processes [Shafi K V, Gedanken A, ProzorovR, Balogh J. Sonochemical preparation and size-dependent properties ofnanostructured CoFe₂O₄ particles. Chemistry of Materials. 1998,10(11):3445-50, incorporated herein by reference in its entirety],microemulsions [Vestal C R, Zhang Z J. Synthesis of CoCrFeO₄nanoparticles using microemulsion methods and size-dependent studies oftheir magnetic properties. Chemistry of materials. 2002, 14(9):3817-22,incorporated herein by reference in its entirety], pulsed laser ablationin liquid [Waag F, Gökce B, Kalapu C, Bendt G. Salamon S, Landers J,Hagemann U, Heidelmann M, Schulz S, Wende H, Hartmann N. Adjusting thecatalytic properties of cobalt ferrite nanoparticles by pulsed laserfragmentation in water with defined energy dose. Scientific Reports.2017, 7(1):13161, incorporated herein by reference in its entirety],ball milling [Khedr M H, Omar A A, Abdel-Moaty S A. Magneticnanocomposites: preparation and characterization of Co-ferritenanoparticles. Colloids and surfaces A: Physicochemical and engineeringaspects. 2006, 281(1-3):8-14, incorporated herein by reference in itsentirety], co-precipitation [Dabagh S, Chaudhary K, Haider Z, Ali J.Study of structural phase transformation and hysteresis behavior ofinverse-spinel α-ferrite nanoparticles synthesized by co-precipitationmethod. Results in Physics. 2018, 8:93-8; and Rani B J, Ravina M,Saravanakumar B, Ravi G, Ganesh V, Ravichandran S, Yuvakkumar R.Ferrimagnetism in cobalt ferrite (CoFe₂O₄) nanoparticles.Nano-Structures & Nano-Objects. 2018, 14:84-91, each incorporated hereinby reference in their entirety], hydrothermal [Zhang W, Zuo X, Zhang D,Wu C, Silva S R. Cr3+ substituted spinel ferrite nanoparticles with highcoercivity. Nanotechnology. 2016, 27(24):245707, incorporated herein byreference in its entirety], sol-gel [Ashour A H, El-Batal A I, AbdelMaksoud M I A, El-Sayyad G S, Labibc S, Abdeltwab E, El-Okr M M.Antimicrobial activity of metal-substituted cobalt ferrite nanoparticlessynthesized by sol-gel technique. Particuology. 2018; volume 40, pages141-151, incorporated herein by reference in its entirety], solvothermal[Kalam A, Al-Sehemi A G, Assiri M, Du G, Ahmad T, Ahmad I, Pannipara M.Modified Solvothermal synthesis of cobalt ferrite (CoFe₂O₄) magneticnanoparticles photocatalysts for degradation of methylene blue withH₂O₂/visible light. Results in Physics. 2018 Jan. 31, incorporatedherein by reference in its entirety], aerosol spray pyrolysis methods[Hong D, Yamada Y, Sheehan M, Shikano S, Kuo C H, Tian M, Tsung C K,Fukuzumi S. Mesoporous nickel ferrites with spinel structure prepared byan aerosol spray pyrolysis method for photocatalytic hydrogen evolution.ACS Sustainable Chemistry & Engineering. 2014, 2(11):2588-94,incorporated herein by reference in its entirety], reverse micelles[Morrison S A, Cahill C L, Carpenter E E, Calvin S, Harris V G.Preparation and characterization of MnZn-ferrite nanoparticles usingreverse micelles. Journal of applied physics. 2003, 93(10):7489-91,incorporated herein by reference in its entirety], and biogenic methodsusing bacteria such as Geobacter sulfurreducens [Céspedes E, Byrne J M.Farrow N, Moise S, Coker V S, Bencsik M, Lloyd J R. Telling N D.Bacterially synthesized ferrite nanoparticles for magnetic hyperthermiaapplications. Nanoscale. 2014, 6(21):12958-70, incorporated herein byreference in its entirety].

Antimicrobial activities of several metal and metal oxide nanoparticlesincluding silver [Ali S G, Ansari M A, Khan H M, Jalal M, Mahdi A A,Cameotra S S. Antibacterial and Antibiofilm Potential of GreenSynthesized Silver Nanoparticles against Imipenem Resistant ClinicalIsolates of P. aeruginosa. BioNanoScience. 2018:1-0, incorporated hereinby reference in its entirety], gold [Payne J N, Waghwani H K, Connor MG, Hamilton W, Tockstein S, Moolani H, Chavda F, Badwaik V, Lawrenz M B,Dakshinamurthy R. Novel synthesis of kanamycin conjugated goldnanoparticles with potent antibacterial activity. Frontiers inmicrobiology. 2016, 7:607, incorporated herein by reference in itsentirety], ZnO [Jalal M, Ansari M A, Ali S G, Khan H M, Rehman S.Anticandidal activity of bioinspired ZnO NPs: effect on growth, cellmorphology and key virulence attributes of Candida species. Artificialcells, nanomedicine, and biotechnology. 2018, 14:1-4, incorporatedherein by reference in its entirety], Al₂O₃[Ansari M A, Khan H M,Alzohairy M A, Jalal M, Ali S G, Pal R, Musarrat J. Green synthesis ofAl₂O₃ nanoparticles and their bactericidal potential against clinicalisolates of multi-drug resistant Pseudomonas aeruginosa. World Journalof Microbiology and Biotechnology. 2015, 31(1):153-64, incorporatedherein by reference in its entirety], Fe₃O₄[Arakha M, Pal S. SamantarraiD, Panigrahi T K, Mallick B C, Pramanik K, Mallick B, Jha S.Antimicrobial activity of iron oxide nanoparticle upon modulation ofnanoparticle-bacteria interface. Scientific reports. 2015, 5:14813; andNehra P, Chauhan R P, Garg N, Verma K. Antibacterial and antifungalactivity of chitosan coated iron oxide nanoparticles. British journal ofbiomedical science. 2018, 75(1):13-8, each incorporated herein byreference in their entirety], and magnetic iron oxide α-Fe₂O₃[Ismail RA, Sulaiman G M, Abdulrahman S A. Marzoog T R. Antibacterial activity ofmagnetic iron oxide nanoparticles synthesized by laser ablation inliquid. Materials Science and Engineering: C. 2015, 53:286-97,incorporated herein by reference in its entirety] nanoparticles havebeen reported. However, very little information is available regardingthe antibacterial properties of transition metal substituted spinelferrite nanoparticles. Recently, Sanpo et al. [Sanpo N, Wen C, Berndt CC, Wang J. Antibacterial properties of spinel ferrite nanoparticles.Microbial pathogens and strategies for combating them: science,technology and education. Spain: Formatex Research Centre. 2013:239-50,incorporated herein by reference in its entirety], Samavati and Ismailet al. [Samavati A, Ismail A F. Antibacterial properties ofcopper-substituted cobalt ferrite nanoparticles synthesized byco-precipitation method. Particuology. 2017; 30:158-63, incorporatedherein by reference in its entirety], and Ashour et al. [Ashour A H,El-Batal A I, Abdel Maksoud M I A, El-Sayyad G S, Labibc S, Abdeltwab E,El-Okr M M. Antimicrobial activity of metal-substituted cobalt ferritenanoparticles synthesized by sol-gel technique. Particuology. 2018;volume 40, pages 141-151, incorporated herein by reference in itsentirety] investigated antibacterial activities of copper, zinc, nickel,and manganese substituted cobalt ferrite nanoparticles for preventingmicrobial infections.

In view of the forgoing, one objective of the present disclosure is toprovide a method of making spinel ferrite nanoparticles comprising achromium-substituted copper ferrite. Another objective of the presentdisclosure is to provide a method of preventing or reducing microbialgrowth on a surface by applying spinel ferrite nanoparticles to thesurface.

BRIEF SUMMARY OF THE INVENTION

According to a first aspect, the present disclosure relates to a methodof making spinel ferrite nanoparticles comprising a chromium-substitutedcopper ferrite of formula (I),CuCr_(x)Fe_(2-x)O₄  (I)wherein x is greater than 0 and smaller than 2. The method involves thesteps of mixing a copper(II) salt, a chromium(III) salt, an iron(III)salt, an inorganic base, and water to form a mixture, heating themixture to form a precipitate, and drying the precipitate, therebyproducing the spinel ferrite nanoparticles,

In one embodiment, the inorganic base is sodium hydroxide.

In one embodiment, the mixture has a pH in a range of 10-12.

In one embodiment, the mixture is heated at a first temperature of40-100° C. for 0.1-3 hours, and subsequently at a second temperature of110-180° C. for 1-6 hours.

In one embodiment, the precipitate is dried at a temperature of 40-100°C. for 1-24 hours.

In one embodiment, the copper(II) salt is copper(II) nitrate.

In one embodiment, the chromium(III) salt is chromium(III) chloride.

In one embodiment, the iron(III) salt is iron(III) nitrate.

In one embodiment, the chromium-substituted copper ferrite of formula(I) is at least one selected from the group consisting ofCuCr_(0.2)Fe_(1.8)O₄, CuCr_(0.4)Fe_(1.6)O₄, CuCr_(0.6)Fe_(1.4)O₄,CuCr_(0.8)Fe_(1.2)O₄, and CuCrFeO₄.

In one embodiment, the spinel ferrite nanoparticles have an averageparticle size in a range of 20-90 nm.

In one embodiment, the spinel ferrite nanoparticles are porous.

In one embodiment, the spinel ferrite nanoparticles have a BET surfacearea in a range of 8-30 m²/g.

In one embodiment, the spinel ferrite nanoparticles have an optical bandgap energy value of 1.0-2.0 eV.

According to a second aspect, the present disclosure relates to a methodfor preventing or reducing growth of a microorganism on a surface. Themethod involves applying spinel ferrite nanoparticles onto the surface,wherein (i) the spinel ferrite nanoparticles comprise achromium-substituted copper ferrite of formula (I)CuCr_(x)Fe_(2-x)O₄  (I)wherein x is greater than 0 and smaller than 2, and (ii) the spinelferrite nanoparticles are in contact with the surface for 0.1-24 hours.

In one embodiment, the spinel ferrite nanoparticles have an averageparticle size in a range of 20-90 nm.

In one embodiment, the spinel ferrite nanoparticles are applied onto thesurface as a suspension comprising a solvent and 50 μg/mL to 100 mg/mLof the spinel ferrite nanoparticles relative to a total volume of thesuspension.

In one embodiment, the solvent comprises water.

In one embodiment, the microorganism is a gram-negative bacterium.

In one embodiment, the gram-negative bacterium is Escherichia coli.

According to a third aspect, the present disclosure relates to spinelferrite nanoparticles, comprising at least one chromium-substitutedcopper ferrite selected from the group consisting ofCuCr_(0.4)Fe_(1.6)O₄, CuCr_(0.6)Fe_(1.4)O₄, CuCr_(0.8)Fe_(1.2)O₄, andCuCrFeO₄.

The foregoing paragraphs have been provided by way of generalintroduction, and are not intended to limit the scope of the followingclaims. The described embodiments, together with further advantages,will be best understood by reference to the following detaileddescription taken in conjunction with the accompanying drawings.

BRIEF DESCRIPTION OF THE DRAWINGS

A more complete appreciation of the disclosure and many of the attendantadvantages thereof will be readily obtained as the same becomes betterunderstood by reference to the following detailed description whenconsidered in connection with the accompanying drawings, wherein:

FIG. 1A shows X-ray diffraction (XRD) patterns of spinel ferritenanoparticles containing CuCrFeO₄.

FIG. 1B shows XRD patterns of spinel ferrite nanoparticles containingCuCr_(0.8)Fe_(1.2)O₄.

FIG. 1C shows XRD patterns of spinel ferrite nanoparticles containingCuCr_(0.6)Fe_(1.4)O₄.

FIG. 1D shows XRD patterns of spinel ferrite nanoparticles containingCuCr_(0.4)Fe_(1.6)O₄.

FIG. 1E shows XRD patterns of spinel ferrite nanoparticles containingCuCr_(0.2)Fe_(1.8)O₄.

FIG. 1F shows XRD patterns of spinel ferrite nanoparticles containingCuFe₂O₄.

FIG. 2 is an overlay of FT-IR spectra of spinel ferrite nanoparticlescontaining CuCrFeO₄ (x=1), CuCr_(0.8)Fe_(1.2)O₄ (x=0.8),CuCr_(0.6)Fe_(1.4)O₄ (x=0.6), CuCr_(0.4)Fe_(1.6)O₄ (x=0.4),CuCr_(0.2)Fe_(1.8)O₄ (x=0.2), and CuFe₂O₄ (x=0.0), respectively.

FIG. 3A is an overlay of diffuse reflectance (DR) spectra of spinelferrite nanoparticles containing CuCrFeO₄ (x=1), CuCr_(0.8)Fe_(1.2)O₄(x=0.8), CuCr_(0.6)Fe_(1.4)O₄ (x=0.6), CuCr_(0.4)Fe_(1.6)O₄ (x=0.4),CuCr_(0.2)Fe_(1.8)O₄ (x=0.2), and CuFe₂O₄ (x=0.0), respectively.

FIG. 3B shows band gap energy values of spinel ferrite nanoparticlescontaining CuCrFeO₄ (x=1), CuCr_(0.8)Fe_(1.2)O₄ (x=0.8),CuCr_(0.6)Fe_(1.4)O₄ (x=0.6), CuCr_(0.4)Fe_(1.6)O₄ (x=0.4),CuCr_(0.2)Fe_(1.8)O₄ (x=0.2), and CuFe₂O₄ (x=0.0), respectively.

FIG. 4 is an overlay of nitrogen physisorption isotherm plots of spinelferrite nanoparticles containing CuCrFeO₄ (x=1), CuCr_(0.8)Fe_(1.2)O₄(x=0.8), CuCr_(0.6)Fe_(1.4)O₄ (x=0.6), CuCr_(0.4)Fe_(1.6)O₄ (x=0.4),CuCr_(0.2)Fe_(1.8)O₄ (x=0.2), and CuFe₂O₄ (x=0.0), respectively.

FIG. 5A shows pore size distribution of spinel ferrite nanoparticlescontaining CuFe₂O₄ (x=0.0).

FIG. 5B shows pore size distribution of spinel ferrite nanoparticlescontaining CuCr_(0.2)Fe_(1.8)O₄ (x=0.2).

FIG. 5C shows pore size distribution of spinel ferrite nanoparticlescontaining CuCr_(0.4)Fe_(1.6)O₄ (x=0.4).

FIG. 5D shows pore size distribution of spinel ferrite nanoparticlescontaining CuCr_(0.6)Fe_(1.4)O₄ (x=0.6).

FIG. 5E shows pore size distribution of spinel ferrite nanoparticlescontaining CuCr_(0.8)Fe_(1.2)O₄ (x=0.8).

FIG. 5F shows pore size distribution of spinel ferrite nanoparticlescontaining CuCrFeO₄ (x=1).

FIG. 6A is a scanning electron microscope (SEM) image of spinel ferritenanoparticles containing CuFe₂O₄ (x=0.0).

FIG. 6B is an SEM image of spinel ferrite nanoparticles containingCuCr_(0.2)Fe_(1.8)O₄ (x=0.2).

FIG. 6C is an SEM image of spinel ferrite nanoparticles containingCuCr_(0.4)Fe_(1.6)O₄ (x=0.4).

FIG. 6D is an SEM image of spinel ferrite nanoparticles containingCuCr_(0.6)Fe_(1.4)O₄ (x=0.6).

FIG. 6E is an SEM image of spinel ferrite nanoparticles containingCuCr_(0.8)Fe_(1.2)O₄ (x=0.8).

FIG. 6F is an SEM image of spinel ferrite nanoparticles containingCuCrFeO₄ (x=1).

FIG. 7A is an SEM image of Escherichia coli cells.

FIG. 7B is an SEM image of Escherichia coli cells treated with spinelferrite nanoparticles containing CuFe₂O₄.

FIG. 7C is an SEM image of Escherichia coli cells treated with spinelferrite nanoparticles containing CuCr_(0.2)Fe_(1.8)O₄.

FIG. 7D is an SEM image of Escherichia coli cells treated with spinelferrite nanoparticles containing CuCr_(0.4)Fe_(1.6)O₄.

FIG. 7E is an SEM image of Escherichia coli cells treated with spinelferrite nanoparticles containing CuCr_(0.6)Fe_(1.4)O₄.

FIG. 7F is an SEM image of Escherichia coli cells treated with spinelferrite nanoparticles containing CuCr_(0.8)Fe_(1.2)O₄.

FIG. 7G is an SEM image of Escherichia coli cells treated with spinelferrite nanoparticles containing CuCrFeO₄.

DETAILED DESCRIPTION OF THE EMBODIMENTS

Embodiments of the present disclosure will now be described more fullyhereinafter with reference to the accompanying drawings, in which some,but not all embodiments of the disclosure are shown.

The present disclosure will be better understood with reference to thefollowing definitions. As used herein, the words “a” and “an” and thelike carry the meaning of “one or more”. Within the description of thisdisclosure, where a numerical limit or range is stated, the endpointsare included unless stated otherwise. Also, all values and subrangeswithin a numerical limit or range are specifically included as ifexplicitly written out.

As used herein, the terms “compound” and “product” are usedinterchangeably, and are intended to refer to a chemical entity, whetherin the solid, liquid or gaseous phase, and whether in a crude mixture orpurified and isolated.

As used herein, a salt refers to an ionic compound derivable from theneutralization reaction of an acid and a base. Salts are composed ofrelated numbers of cations (positively charged ions) and anions(negatively charged ions) such that the product is electrically neutral(without a net charge). These component ions may be inorganic (e.g.chloride, Cl⁻) or organic (e.g. acetate, CH₃CO₂ ⁻) and may be monoatomic(e.g. fluoride, F) or polyatomic (e.g. sulfate, SO₄ ²⁺). Exemplaryconventional salts include, but are not limited to, those derived frominorganic acids including, but not limited to, hydrochloric,hydrobromic, sulfuric, sulfamic, phosphoric, and nitric; and thosederived from organic acids including, but not limited to, acetic,propionic, succinic, glycolic, stearic, lactic, malic, tartaric, citric,ascorbic, pamoic, maleic, hydroxymaleic, phenylacetic, glutamic,benzoic, salicyclic, sulfanilic, 2-acetoxybenzoic, fumaric,toluenesulfonic, methanesulfonic, ethane disulfonic, oxalic, isethionic,and mixtures and hydrates thereof and the like. The present disclosureincludes all hydration states of a given salt or formula, unlessotherwise noted. For example, copper(II) nitrate includes anhydrousCu(NO₃)₂, monohydrate Cu(NO₃)₂·H₂O, hemi(pentahydrate) Cu(NO₃)₂·2.5H₂O,trihydrate Cu(NO₃)₂·3H₂O, and any other hydrated forms or mixtures.Chromium(III) chloride includes anhydrous CrCl₃, and hydrated forms suchas chromium trichloride hexahydrate CrCl₃·6H₂O. Iron(III) nitrateincludes anhydrous Fe(NO₃)₃, and hydrated forms such as iron(III)nitrate nonahydrate Fe(NO₃)₃·9H₂O.

The present disclosure is intended to include all isotopes of atomsoccurring in the present compounds. Isotopes include those atoms havingthe same atomic number but different mass numbers. By way of generalexample, and without limitation, isotopes of hydrogen include deuteriumand tritium, isotopes of carbon include ¹³C and ¹⁴C, isotopes of oxygeninclude ¹⁶O, ¹⁷, and ¹⁸O, isotopes of copper include ⁶³Cu and ⁶⁵Cu,isotopes of chromium include ⁵⁰Cr, ⁵²Cr, ⁵³Cr, and ⁵⁴Cr, and isotopes ofiron include ⁵⁴Fe, ⁵⁶Fe, ⁵⁷Fe, and ⁵⁸Fe. Isotopically labeled compoundsof the disclosure can generally be prepared by conventional techniquesknown to those skilled in the art or by processes and methods analogousto those described herein, using an appropriate isotopically labeledreagent in place of the non-labeled reagent otherwise employed.

A first aspect of the present disclosure relates to a method of makingspinel ferrite nanoparticles comprising a chromium-substituted copperferrite of formula (I),CuCr_(x)Fe_(2-x)O₄  (I)wherein x is greater than 0 and smaller than 2, preferably x is in arange of 0<x≤1.5, more preferably x is in a range of 0.1≤x≤1.0. Themethod involves the steps of mixing a copper(II) salt, a chromium(III)salt, an iron(III) salt, an inorganic base, and water to form a mixture,heating the mixture to form a precipitate, and drying the precipitate,thereby producing the spinel ferrite nanoparticles.

As defined herein, a spinel is a metal oxide compound with a generalformula A²⁺B₂ ³⁺O₄ ²⁻, where “A” and “B” are metal ions. In oneembodiment, “A” may be Zn, Cu, Co, Mn, Ni, Mg, Be, and/or Ti, and “B”may be Al, Fe, Cr, and/or V. Preferably, spinel compounds are in theform of crystals, with the oxide anions arranged in a cubic close-packedlattice, and with the metal ions occupying octahedral and/or tetrahedralsites within the lattice. Preferably, the A²⁺ metal ions occupy thetetrahedral sites, and the B³⁺ metal ions occupy the octahedral sites,though there may be instances where the metal ions are switched. The A²⁺and B³⁺ metal ions may occupy sites in the lattice at regular spacingsor may be distributed randomly. A spinel ferrite is defined herein as aniron-containing spinel compound with a formula C²⁺D_(2-y) ³⁺Fe_(y)O₄ ²⁻,where “C” and “D” are metal ions, and “y” is in a range of 0≤y≤2. In oneembodiment, “C” may be Zn, Cu, Co, Mn, Ni, Mg, Be, and/or Ti, and “D”may be Al, Fe, Cr, and/or V. Preferably, the tetrahedral sites of aspinel ferrite are occupied by C²⁺ metal ions, and the octahedral sitesare occupied by Fe³⁺ ions and substitution D³⁺ metal ions.

In one or more embodiments, the spinel ferrite disclosed hereincomprises a chromium-substituted copper ferrite of formula (I),CuCr_(x)Fe_(2-x)O₄  (I)wherein x is greater than 0 and smaller than 2, preferably x is in arange of 0<x≤1.5, more preferably x is in a range of 0.1≤x≤1.0. Forexample, x may be 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.55,0.6, 0.65, 0.7, 0.75, 0.8, 0.85, 0.9, 0.95, or 1. Alternatively, x maybe 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, or 1.9. In preferredembodiments, x is 0.2, 0.4, 0.6, 0.8, or 1. In a related embodiment, thespinel ferrite disclosed herein comprises a chromium-substituted copperferrite of formula (I) which is at least one selected from the groupconsisting of CuCr_(0.2)Fe_(1.8)O₄, CuCr_(0.4)Fe_(1.6)O₄,CuCr_(0.6)Fe_(1.4)O₄, CuCr_(0.8)Fe_(1.2)O₄, and CuCrFeO₄. Atomic ratiosof the chromium-substituted copper ferrites may be determined byelemental analysis techniques such as energy-dispersive X-rayspectroscopy (EDX), X-ray photoelectron spectroscopy (XPS), inductivelycoupled plasma mass spectrometry (ICP-MS), and neutron activationanalysis. In at least one embodiment, the spinel ferrite disclosedherein consists essentially of the chromium-substituted copper ferriteof formula (I) (i.e. oxygen atoms and metal atoms including copper,chromium, and iron), and is devoid of other metal atoms such as zinc,nickel, cobalt, aluminum, and manganese.

A particle is defined as a small object that behaves as a whole unitwith respect to its transport and properties. The spinel ferrite of thepresent disclosure in any of its embodiments may be in the form ofparticles of the same shape or different shapes, and of the same size ordifferent sizes. An average diameter (e.g., average particle size) ofthe particle, as used herein, refers to the average linear distancemeasured from one point on the particle through the center of theparticle to a point directly across from it. Microparticles areparticles having an average diameter between 0.1 and 100 μm in size.Nanoparticles are particles having an average diameter between 1 and 100nm in size.

In one or more embodiments, the spinel ferrite disclosed herein is inthe form of nanoparticles. The exceptionally high surface area to volumeratio of nanoparticles may cause the nanoparticles to exhibitsignificantly different or even novel properties from those observed inindividual atoms/molecules, fine particles and/or bulk materials. In apreferred embodiment, the spinel ferrite described herein is in the formof nanoparticles, which are spherical or substantially spherical (e.g.oval, oblong, etc.) in shape. In another preferred embodiment, thespinel ferrite described herein is in the form of nanoparticles, whichare angular shaped (e.g. rectangles, triangles, pentagons, prisms,prismoids, etc.). Alternatively, it is envisaged that the spinel ferritenanoparticles may have an irregular shape. However, the spinel ferritenanoparticles described herein may have various shapes other thanspherical or angular shape and may be of any shape that provides desiredantimicrobial activity. For example, the spinel ferrite nanoparticles ofthe present disclosure may demonstrate a variety of morphologiesincluding, but not limited to, nanosheets, nanoplatelets, nanocrystals,nanospheres, nanohexagons, nanodisks, nanocubes, nanowires, nanofibers,nanoribbons, nanorods, nanotubes, nanocylinders, nanogranules,nanowhiskers, nanoflakes, nanofoils, nanopowders, nanoboxes, nanostars,tetrapods, nanobelts, nanaourchins, nanofloweres, and mixtures thereof.

In one or more embodiments, the spinel ferrite nanoparticles have anaverage particle size in a range of 20-99 nm, 25-95 nm, 30-90 nm, 35-85nm, 40-80 nm, 45-75 nm, 50-70 nm, or 55-65 nm. However, in certainembodiments, the spinel ferrite disclosed herein has an average particlesize smaller than 20 nm or greater than 99 nm. For example, the spinelferrite disclosed herein may be in the form of microparticles having anaverage particle size of 0.1-100 μm, 0.5-90 μm, 1-80 μm, 5-70 μm, 10-60μm. 20-50 μm, or 30-40 μm. In one embodiment, the spinel ferritenanoparticles may be clustered together as agglomerates having anaverage diameter in a range of 0.1-5 μm, 0.2-4 μm, 0.4-2 μm, or 0.5-1μm. In a preferred embodiment, the spinel ferrite nanoparticles are wellseparated from one another and do not form agglomerates. The size andshape of particles may be analyzed by techniques such as dynamic lightscattering (DLS), scanning electron microscopy (SEM), transmissionelectron microscopy (TEM), and/or atomic force microscopy (AFM).

As used herein, “dispersity” is a measure of the heterogeneity of sizesof molecules or particles in a mixture. In probability theory orstatistics, the coefficient of variation (CV), also known as relativestandard deviation (RSD) is a standardized measure of dispersion of aprobability distribution. It is expressed as a percentage and may bedefined as the ratio of the standard deviation (a) to the mean (p, orits absolute value |μ|). The coefficient of variation or relativestandard deviation is widely used to express precision and/orrepeatability. It may show the extent of variability in relation to themean of a population. In a preferred embodiment, the spinel ferritenanoparticles of the present disclosure have a narrow size dispersion,i.e. are monodisperse. As used herein, “monodisperse”, “monodispersed”,and/or “monodispersity” refers to spinel ferrite nanoparticles whichhave a CV or RSD of less than 30%, preferably less than 25%, preferablyless than 20%, preferably less than 15%, preferably less than 12%,preferably less than 10%, preferably less than 8%, preferably less than5%.

In one or more embodiments, the spinel ferrite nanoparticles are porous.The term “microporous” means the pores of the particles have an averagepore size of less than 2 nm. The term “mesoporous” means the pores ofthe particles have an average pore size of 2-50 nm. The term“macroporous” means the pores of the particles have an average pore sizelarger than 50 nm. Pore size may be determined by methods including, butnot limited to, gas adsorption (e.g. N₂ adsorption), mercury intrusionporosimetry, and imaging techniques such as scanning electron microscopy(SEM), and x-ray computed tomography (XRCT). The spinel ferritenanoparticles may be mesoporous, or microporous. In one embodiment, thespinel ferrite nanoparticles are microporous, and have an average poresize in a range of 0.05-1.99 nm, 0.1-1.9 nm, 0.2-1.8 nm, 0.3-1.7 nm,0.4-1.6 nm, 0.5-1.5 nm, 0.6-1.4 nm. 0.7-1.3 nm, 0.8-1.2 nm, or 0.9-1.1nm. In another embodiment, the spinel ferrite nanoparticles aremesoporous, and have an average pore size in a range of 2-15 nm, 3-14nm, 4-13 nm, 5-12 nm, 6-11 nm, 7-10 nm, or 8-9 nm. In a preferredembodiment, the spinel ferrite nanoparticles comprising thechromium-substituted copper ferrite in any of their embodiments have anaverage pore size that is at least 1.5 nm greater than the average poresize of substantially similar spinel ferrite nanoparticles comprising acopper ferrite lacking chromium substitution (e.g. CuFe₂O₄), preferablyat least 2 nm greater, preferably at least 3 nm greater, preferably atleast 4 nm greater, preferably at least 5 nm greater, preferably atleast 6 nm greater, preferably at least 7 nm greater, preferably atleast 8 nm greater than the average pore size of substantially similarspinel ferrite nanoparticles comprising a copper ferrite lackingchromium substitution (see Table 2).

The Brunauer-Emmet-Teller (BET) theory (S. Brunauer, P. H. Emmett, E.Teller, J. Am. Chem. Soc. 1938, 60, 309-319, incorporated herein byreference) aims to explain the physical adsorption of gas molecules on asolid surface and serves as the basis for an important analysistechnique for the measurement of a specific surface area of a material.Specific surface area is a property of solids which is the total surfacearea of a material per unit of mass, solid or bulk volume, or crosssectional area. In most embodiments, pore volume and BET surface areaare measured by gas adsorption analysis, preferably N₂ adsorptionanalysis. In one or more embodiments, the spinel ferrite nanoparticlesof the present disclosure have a BET surface area in a range of 2-50m²/g, preferably 4-40 m²/g, preferably 8-30 m²/g, preferably 10-25 m²/g,preferably 11-20 m²/g, preferably 12-19 m²/g, preferably 13-18 m²/g,preferably 14-17 m²/g, preferably 15-16 m²/g. In one embodiment, thespinel ferrite nanoparticles, comprising the chromium-substituted copperferrite of formula CuCr_(x)Fe_(2-x)O₄, have a smaller BET surface areawhen the chromium content is higher (i.e. surface area decreases as “x”increases from 0.2 to 1) (see Table 2). In a preferred embodiment, thespinel ferrite nanoparticles comprising the chromium-substituted copperferrite in any of their embodiments have a BET surface area that is atleast 0.5 m²/g smaller than the average pore size of substantiallysimilar spinel ferrite nanoparticles comprising a copper ferrite lackingchromium substitution (e.g. CuFe₂O₄), preferably at least 1 m²/gsmaller, preferably at least 2 m²/g smaller, preferably at least 3 m²/gsmaller, preferably at least 4 m²/g smaller, preferably at least 5 m²/gsmaller, preferably at least 6 m²/g smaller, preferably at least 7 m²/gsmaller, preferably at least 8 m²/g smaller, preferably at least 9 m²/gsmaller, preferably at least 10 m²/g smaller than the BET surface areaof substantially similar spinel ferrite nanoparticles comprising acopper ferrite lacking chromium substitution (e.g. CuFe₂O₄) (see Table2).

As used herein, band gap energy, band gap, and/or energy gap refers toan energy range in a solid where no electron states can exist. In graphsof the electronic band structure of solids, the band gap generallyrefers to the energy difference (in electron volts) between the top ofthe valence band and the bottom of the conduction band in insulatorsand/or semiconductors. It is generally the energy required to promote avalence electron bound to an atom to become a conduction electron, whichis free to move within the crystal lattice and serve as a charge carrierto conduct electric current. Band gap energies for the spinel ferritenanoparticles described herein may be obtained using opticalspectroscopies, e.g. UV-vis spectroscopy and/or electrochemicalmeasurements, e.g. cyclic voltammetry (CV) and differential pulsevoltammetry (DPV). In one or more embodiments, the spinel ferritenanoparticles described herein in any of their embodiments have a bandgap energy of 1.0-2.5 eV, 1.1-2.0 eV, 1.2-1.9 eV, 1.3-1.8 eV, 1.4-1.7eV, or 1.5-1.6 eV. In certain embodiments, the spinel ferritenanoparticles have a band gap energy smaller than 1.0 eV or greater than2.5 eV.

As used herein. “crystallite size”, “crystalline size” and/or “crystalsize” refers to a Scherrer derived particle size or crystal size. AScherrer derived particle size or crystal size relates the mean (volumeaverage) crystal or particle size of a powder to the broadening of itspowder diffraction peaks. Crystallite size is different than theaforementioned particle size as a particle may be made up of severalindividual crystallites. The spinel ferrite nanoparticles may becrystalline, polycrystalline, or amorphous. Preferably, the spinelferrite nanoparticles are crystalline. In a preferred embodiment, thespinel ferrite nanoparticles of the present disclosure in any of theirembodiments have an average crystallite size of 15-45 nm, preferably18-40 nm, preferably 20-38 nm, preferably 23-35 nm, preferably 25-32 nm,preferably 28-30 nm. Crystallite size of the spinel ferritenanoparticles may be determined via X-ray diffraction (XRD) technique.In a preferred embodiment, the spinel ferrite nanoparticles comprisingthe chromium-substituted copper ferrite in any of their embodiments havean average crystallite size that is at least 6 nm smaller than theaverage crystallite size of substantially similar spinel ferritenanoparticles comprising a copper ferrite lacking chromium substitution(e.g. CuFe₂O₄), preferably at least 9 nm smaller, preferably at least 12nm smaller, preferably at least 15 nm smaller, preferably at least 18 nmsmaller, preferably at least 20 nm smaller, preferably at least 23 nmsmaller, preferably at least 25 nm smaller, preferably at least 30 nmsmaller than the average crystallite size of substantially similarspinel ferrite nanoparticles comprising a copper ferrite lackingchromium substitution (e.g. CuFe₂O₄) (see Table 1).

The method of producing the spinel ferrite nanoparticles involves mixinga copper(II) salt, a chromium(III) salt, an iron(III) salt, an inorganicbase, and water to form a mixture. The water may be tap water, distilledwater, bidistilled water, deionized water, deionized distilled water,reverse osmosis water, and/or some other water. In one embodiment, thewater is bidistilled to eliminate trace metals. Preferably the water isdistilled water. In certain embodiments, other solvents including, butnot limited to, alcohols (e.g. methanol, ethanol, n-propanol,i-propanol, n-butanol), and acetone may be used in addition to or inlieu of water.

Prior to the mixing step, the aforementioned reagents (i.e. copper(II),chromium(III), and iron(III) salts, and the inorganic base) may bedissolved in water separately to form respective solutions, which arethen mixed to form the mixture. In an alternative embodiment, the metalsalts (i.e. copper(II), chromium(III), and iron(III) salts) aredissolved in water to form a first mixture, and an aqueous solution ofthe inorganic base is mixed with the first mixture to form the mixture.The mixing may occur via stirring, shaking, sonicating, blending, or byotherwise agitating the mixture. In a preferred embodiment, the mixtureis stirred at a temperature of 50-95° C., 60-90° C., or 70-80° C. for0.1-6 hours, 0.25-3 hours, or 0.5-2 hours. The stirring may by performedby a magnetic stirrer or an overhead stirrer. In another embodiment, themixture is left to stand (i.e. not stirred). An external heat source,such as a water bath or an oil bath, an oven, or a heating mantle, maybe employed to heat the mixture.

In one or more embodiments, a molar ratio of the copper(II) salt to atotal mole of the chromium(III) salt and the iron(III) salt is in arange of 1:1 to 1:4, preferably 2:3 to 1:3, more preferably 1:1.8 to2:5, or about 1:2. Based on the aforementioned chromium-substitutedcopper ferrite of formula CuCr_(x)Fe_(2-x)O₄, a molar ratio of thechromium(III) salt to the iron(III) salt may be represented as a ratioof “x” to “2−x”. For example, a molar ratio of the chromium(III) salt tothe iron(III) salt may be about 1:19 for the preparation of spinelferrite nanoparticles comprising chromium-substituted copper ferrite offormula CuCr_(0.1)Fe_(1.9)O₄. A molar ratio of the chromium(III) salt tothe iron(III) salt may be about 1:1 for the preparation of spinelferrite nanoparticles comprising chromium-substituted copper ferrite offormula CuCrFeO₄. In one embodiment, an overall concentration of thecopper(II) salt, the chromium(III) salt and the iron(III) salt in themixture may be in the range of 0.01-50 M, 0.05-40 M, 0.1-30 M, 0.5-15 M,1-10 M, 2-6 M, or 3-4 M.

Non-limiting examples of the copper(II) salt include copper(II) nitrate,copper(II) chloride, copper(II) sulfate, copper(II) bromide, copper(II)iodide, and mixtures thereof. The copper(II) salt used herein may be inany hydration state, for instance, copper(II) nitrate includes, withoutlimitation, Cu(NO₃)₂, Cu(NO₃)₂·H₂O, Cu(NO₃)₂·2.5H₂O, Cu(NO₃)₂·3H₂O, andCu(NO₃)₂·6H₂O. In certain embodiments, a copper salt having a differentoxidation state, such as +1, may be used in addition to or in lieu ofthe copper(II) salt. In a preferred embodiment, the copper(II) salt iscopper(II) nitrate.

Non-limiting examples of the chromium(III) salt include chromium(III)chloride, chromium(III) nitrate, chromium(III) sulfate, chromium(III)bromide, chromium(III) fluoride, and mixtures thereof. The chromium(III)salt used herein may be in any hydration state, for example,chromium(III) chloride includes, but are not limited to, CrCl₃,CrCl₃.5H₂O, and CrCl₃.6H₂O. In certain embodiments, a chromium salthaving a different oxidation state, such as +2, may be used in additionto or in lieu of the chromium(III) salt. In a preferred embodiment, thechromium(III) salt is chromium(III) chloride.

Non-limiting examples of the iron(III) salt include iron(III) nitrate,iron(III) chloride, iron(III) sulfate, iron(III) bromide, iron(III)fluoride, iron(III) phosphate, and mixtures thereof. The iron(III) saltused herein may be in any hydration state, for instance, iron(III)nitrate includes, without limitation, Fe(NO₃)₃, Fe(NO₃)₃·6H₂O, andFe(NO₃)₃·9H₂O. In certain embodiments, an iron salt having a differentoxidation state, such as +2, may be used in addition to or in lieu ofthe iron(III) salt. In a preferred embodiment, the iron(III) salt isiron(III) nitrate.

In one embodiment, the inorganic base is sodium hydroxide (NaOH),potassium hydroxide (KOH), ammonium hydroxide (NH₄OH), cesium hydroxide(CsOH), lithium hydroxide (LiOH), calcium hydroxide (Ca(OH)₂), bariumhydroxide (Ba(OH)₂), strontium hydroxide (Sr(OH)₂), sodium carbonate(Na₂CO₃), potassium carbonate (K₂CO₃), cesium carbonate (Cs₂CO₃),trisodium phosphate (Na₃PO₄), or a mixture thereof. In a preferredembodiment, the inorganic base is sodium hydroxide. In a preferredembodiment, the pH of the mixture is maintained at a range of 9-13,preferably 9.5-12.5, preferably 10-12, preferably 10.5-11.5, or about11. The pH of the mixture may be monitored using a pH meter, pH testpapers, and/or pH indicators.

The method also involves the step of heating the mixture to form aprecipitate. In a preferred embodiment, the mixture is heated at a firsttemperature of 40-100° C., preferably 50-90° C., preferably 60-85° C.,preferably 70-80° C. for 0.1-3 hours, 0.25-2 hours, 0.5-1 hour, or about40 minutes, and subsequently at a second temperature of 110-180° C.,preferably 120-170° C., preferably 130-160° C., preferably 140-150° C.for 1-6 hours, 2-5 hours, 2.5-4 hours, or about 3 hours. A precipitationmay be formed during the heating processes and be separated (e.g.filtered off, centrifuged) from the aforementioned mixture.Alternatively, the mixture may be heated in a single stage. For example,the mixture may be heated at a temperature of 40-180° C., preferably60-160° C., more preferably 80-130° C. for 0.5-12 hours, 1-6 hours, or2-4 hours.

The method further involves the step of drying the precipitate at atemperature of 40-600° C., 60-500° C. 80-400° C., 100-300° C., or150-250° C. for 1-48 hours, 2-36 hours, 4-24 hours, or 8-12 hours toproduce the spinel ferrite nanoparticles. In one embodiment, this stepinvolves an initial heating of the precipitate at a temperature of40-150° C., preferably 50-120° C., more preferably 60-90° C., or about70° C. for 1-36 hours, 2-24 hours, 4-12 hours, or about 6 hours, and anadditional heat treatment (i.e. annealing) of the precipitate at atemperature of 200-600° C., preferably 300-500° C., more preferably350-450° C., or about 400° C. for 1-24 hours, 2-12 hours, 3-6 hours, orabout 4 hours. In a related embodiment, the initial heating may beperformed using a hot plate, an oven, or in some embodiments, theprecipitate may be subjected to a vacuum, or a rotary evaporator. Inanother related embodiment, the additional heat treatment (i.e.annealing) may be conducted in air within an oven or furnace. Also, insome embodiments, the precipitate may not be annealed via additionalheating in air, but in oxygen-enriched air, an inert gas, or a vacuum.

In at least one embodiment, the currently disclosed method in any of itsembodiments does not involve the usage of chelating agent such as urea,thiourea, citric acid, ethylenediaminetetraacetic acid (EDTA), oxalicacid, malic acid, sebacic acid, tartaric acid, glucose, amino acids suchas glutamine and histidine, as well as other triprotic acids such asisocitric acid, aconitic acid, and propane-1,2,3-tricarboxylic acid,which are commonly applied in sol-gel auto-combustion technique for theproduction of spinel nanoparticles.

A further aspect of the present disclosure relates to spinel ferritenanoparticles, comprising at least one chromium-substituted copperferrite selected from the group consisting of CuCr_(0.4)Fe_(1.6)O₄,CuCr_(0.6)Fe_(1.4)O₄, CuCr_(0.8)Fe_(1.2)O₄, and CuCrFeO₄. In oneembodiment, the spinel ferrite nanoparticles comprising at least onechromium-substituted copper ferrite selected from the group consistingof CuCr_(0.4)Fe_(1.6)O₄, CuCr_(0.6)Fe_(1.4)O₄, CuCr_(0.8)Fe_(1.2)O₄, andCuCrFeO₄ are made by the method of the first aspect of the currentdisclosure. Thus, the spinel ferrite nanoparticles may have similarproperties as described for those in the first aspect, such as averageparticle size, surface area, pore size, crystallite size, and/or someother property. Preferably, the spinel ferrite nanoparticles of thecurrent aspect have aforementioned composition ratios, shapes, averageparticle sizes, BET surface areas, average pore sizes, and averagecrystallite sizes. In one embodiment, these spinel ferrite nanoparticlesmay have an average particle size in a range of 20-80 nm. 25-75 nm,30-70 nm, 35-65 nm, 40-60 nm, or 45-55 nm. In another embodiment, thesespinel ferrite nanoparticles have an average pore size in a range of5-20 nm, 6-19 nm, 7-18 nm, 8-17 nm, 9-16 nm, 10-15 nm. 11-14 nm, or12-13 nm. In another embodiment, these spinel ferrite nanoparticles havea BET surface area in a range of 5-18 m²/g, preferably 6-17 m²/g,preferably 7-16 m²/g, preferably 8-15 m²/g, preferably 9-14 m²/g,preferably 10-13 m²/g, preferably 11-12 m²/g. In another embodiment,these spinel ferrite nanoparticles have an average crystallite size in arange of 15-40 nm, preferably 18-38 nm, preferably 20-35 nm, preferably23-32 nm, preferably 25-30 nm, preferably 26-28 nm. Alternatively, thespinel ferrite nanoparticles comprising at least onechromium-substituted copper ferrite selected from the group consistingof CuCr_(0.4)Fe_(1.6)O₄, CuCr_(0.6)Fe_(1.4)O₄, CuCr_(0.8)Fe_(1.2), andCuCrFeO₄ may be prepared by other applicable processes including, butnot limited to, sol-gel method, sol-gel auto-combustion, sonochemicalmethod, ball milling, hydrothermal process, solvothermal process,aerosol spray pyrolysis method, and biological approach.

According to a second aspect, the present disclosure relates to a methodfor preventing or reducing growth of a microorganism on a surface usingspinel ferrite nanoparticles comprising chromium-substituted copperferrite of formula CuCr_(x)Fe_(2-x)O₄, wherein x is greater than 0 andsmaller than 2, preferably x is in a range of 0<x≤1.5, more preferably xis in a range of 0.1≤x≤1.0. In one or more embodiments, the spinelferrite used herein comprises a chromium-substituted copper ferritewhich is at least one selected from the group consisting ofCuCr_(0.2)Fe_(1.8)O₄, CuCr_(0.4)Fe_(1.6)O₄, CuCr_(0.6)Fe_(1.4)O₄,CuCr_(0.8)Fe_(1.2)O₄, and CuCrFeO₄. Preferably, these spinel ferritenanoparticles may have an average particle size in a range of 20-99 nm,25-95 nm, 30-90 nm, 35-85 nm, 40-80 nm, 45-75 nm, 50-70 nm, or 55-65 nm.The spinel ferrite nanoparticles may have similar properties asdescribed for those in the first aspect, such as average particle size,surface area, pore size, crystallite size, and/or some other property.In preferred embodiments, the spinel ferrite nanoparticles used hereinfor preventing or reducing growth of a microorganism on a surface haveaforementioned composition ratios, shapes, average particle sizes, BETsurface areas, average pore sizes, and average crystallite sizes.

In other embodiments, the spinel ferrite nanoparticles having one ormore dissimilar properties as compared to those described in the firstaspect may be used herein for preventing or reducing growth of amicroorganism on a surface. For example, spinel ferrite nanoparticlesmay be used which have an average particle size smaller than 20 nm orgreater than 99 nm. Alternatively, spinel ferrite nanoparticles may beused which have a BET surface area smaller than 2 m²/g or greater than50 m²/g, and/or an average crystallite size smaller than 15 nm orgreater than 45 nm. In a related embodiment, spinel ferritenanoparticles which are microporous, mesoporous, or macroporous may beused herein for preventing or reducing growth of a microorganism on asurface, or in some embodiments, spinel ferrite nanoparticles which aredense (i.e. non-porous) may be used instead. These spinel ferritenanoparticles with dissimilar properties may be formed by changing theaforementioned reaction conditions, such as solvent, reaction time, pH,and/or temperature. Alternatively, these spinel ferrite nanoparticleswith dissimilar properties may be prepared via different syntheticroutes such as sol-gel auto-combustion, microwave-assisted method,solid-state reaction, and hydrothermal synthesis.

The current method for preventing or reducing growth of a microorganismon a surface involves applying the spinel ferrite nanoparticles onto thesurface. Preferably, the spinel ferrite nanoparticles are in contactwith the surface for 0.1-48 hours, 0.5-36 hours, 1-24 hours, 2-12 hours,or 3-6 hours.

In one or more embodiments, the spinel ferrite nanoparticles are appliedonto the surface as a mixture (e.g. a suspension, a solution, a colloid)comprising the spinel ferrite nanoparticles. Preferably, the spinelferrite nanoparticles are applied onto the surface as a suspensioncomprising a solvent and the spinel ferrite nanoparticles. Thesuspension may comprise 10 μg/mL to 1,000 mg/mL of the spinel ferritenanoparticles relative to a total volume of the suspension, preferably50 μg/mL to 800 mg/mL, preferably 100 μg/mL to 600 mg/mL, preferably 500μg/mL to 400 mg/mL, preferably 1 to 200 mg/mL, preferably 2.5 to 150mg/mL, preferably 4 to 125 mg/mL, preferably 8 to 100 mg/mL, preferably16 to 90 mg/mL, preferably 32 to 80 mg/mL, preferably 40 to 70 mg/mL,preferably 50 to 60 mg/mL.

In one or more embodiments, the spinel ferrite nanoparticles used hereincomprise a chromium-substituted copper ferrite which is at least oneselected from the group consisting of CuCr_(0.2)Fe_(1.8)O₄,CuCr_(0.4)Fe_(1.6)O₄, CuCr_(0.6)Fe_(1.4)O₄, CuCr_(0.8)Fe_(1.2)O₄, andCuCrFeO₄. In a preferred embodiment, the spinel ferrite nanoparticlesused herein comprise a chromium-substituted copper ferrite which is atleast one selected from the group consisting of CuCr_(0.4)Fe_(1.6)O₄,CuCr_(0.8)Fe_(1.2)O₄, and CuCrFeO₄. In one or more embodiments, thesolvent comprises water. In a related embodiment, other compatiblesolvents, such as phosphate-buffered saline, alcohols (e.g. methanol,ethanol, trifluoroethanol, n-propanol, i-propanol, n-butanol, i-butanol,t-butanol, n-pentanol, i-pentanol, 2-methyl-2-butanol,2-trifluoromethyl-2-propanol, 2,3-dimethyl-2-butanol, 3-pentanol,3-methyl-3-pentanol, 2-methyl-3-pentanol, 2-methyl-2-pentanol,2,3-dimethyl-3-pentanol, 3-ethyl-3-pentanol, 2-methyl-2-hexanol,3-hexanol, cyclopropylmethanol, cyclopropanol, cyclobutanol,cyclopentanol, cyclohexanol), acetonitrile, and dimethyl sulfoxide(DMSO), may be used in addition to or in lieu of water. Alternatively,the solvent may be chlorinated solvents (e.g. chlorobenzene,dichloromethane, 1,2-dichloroethane, 1,1-dichloroethane, chloroform),ester solvents (e.g. ethyl acetate, propyl acetate), ethers (e.g.diethyl ether, tetrahydrofuran, 1,4-dioxane, tetrahydropyran, t-butylmethyl ether, cyclopentyl methyl ether, di-iso-propyl ether), aromaticsolvents (e.g. benzene, o-xylene, m-xylene, p-xylene, mixtures ofxylenes, toluene, mesitylene, anisole, 1,2-dimethoxybenzene,α,α,α-trifluoromethylbenzene, fluorobenzene), or a mixture thereof.

As used herein, “microorganism” or “microbe” refers to in particularfungi, and gram-positive and gram-negative bacteria. The term“antimicrobial product” refers to a product demonstrating the capabilityto inhibit or prevent the proliferation of microorganisms. Gram-negativebacteria are bacteria that do not retain the crystal violet stain usedin the gram-staining method of bacterial differentiation. Gram-negativebacteria are considered great medical challenges as the thick outermembrane protects these bacteria from many antibiotics, dyes, anddetergents.

In one or more embodiments, the spinel ferrite nanoparticles are appliedonto the surface as an antimicrobial product containing the spinelferrite nanoparticles at an amount of 0.01-99 wt %, 0.5-95 wt %, 1-90 wt%, 2-80 wt %, 5-70 wt %, 10-60 wt %, 20-50 wt %, 30-40 wt % relative toa total weight of the antibacterial product. In one or more embodiments,the spinel ferrite nanoparticles used herein comprise achromium-substituted copper ferrite which is at least one selected fromthe group consisting of CuCr_(0.2)Fe_(1.8)O₄, CuCr_(0.4)Fe_(1.6)O₄,CuCr_(0.6)Fe_(1.4)O₄, CuCr_(0.8)Fe_(1.2)O₄, and CuCrFeO₄. In a preferredembodiment, the spinel ferrite nanoparticles used herein comprise achromium-substituted copper ferrite which is at least one selected fromthe group consisting of CuCr_(0.4)Fe_(1.6)O₄, CuCr_(0.8)Fe_(1.2)O₄, andCuCrFeO₄.

Exemplary antimicrobial products include, but are not limited to,antimicrobial coatings, hand sanitizer (including rinse off and leave-onand aqueous-based hand disinfectants), preoperative skin disinfectant,bar soap, liquid soap (e.g., hand soap), hospital disinfectants,disinfecting spray solution, household cleansing wipes, surfacesanitizer, personal care disinfecting wipes, body wash, acne treatmentproducts, antibacterial diaper rash cream, antibacterial skin cream,deodorant, antimicrobial creams, topical cream, a wound care item, suchas wound healing ointments, creams, and lotions.

The method disclosed herein may be used to prevent or reduce growth of amicroorganism on the skin of a subject. In a preferred embodiment, thespinel ferrite nanoparticles are applied onto the skin of a subject asan antimicrobial cream comprising 0.01-50 wt %, 0.1-40 wt %, 1-30 wt %,2-20 wt %. 4-15 wt %, or 5-10 wt % of spinel ferrite nanoparticlesrelative to a total weight of the antimicrobial cream. The subject maybe a mammal, such as a human; a non-human primate, such as a chimpanzee,and other apes and monkey species; a farm animal, such as a cow, ahorse, a sheep, a goat, and a pig; a domestic animal, such as a rabbit,a dog, and a cat; a laboratory animal including a rodent, such as a rat,a mouse, and a guinea pig, and the like. The antimicrobial cream mayfurther comprise other formulating components such as occlusioncomponents (e.g. petrolatum), film forming agents (e.g.polyvinylpyrrolidone), thickening agents (e.g. xanthan gum,polyacrylamide polymers), and/or emulsifiers (e.g. fatty acids, fattyalcohols). The formulation techniques of topical creams are generallyknown to those skilled in the art.

Other surfaces suitable for the method disclosed herein include bothhard and soft surfaces. The term “hard surface” includes, but is notlimited to, bathroom surfaces (tub and tile, fixtures, ceramics),kitchen surfaces, countertops, appliances, flooring, glass, automobiles,and the like. “Soft surfaces” include but are not limited to fabrics,leather, carpets, furniture, upholstery and other suitable softsurfaces. The presently disclosed method may also be viable forsanitizing surfaces related to hospital and nursing care facilities andequipment such as hospital room, toilet, beds, sheets, pillows,wheelchairs, and canes.

The method disclosed herein may also be used to prevent or reduce growthof a microorganism on the surface of an artificial restoration ormedical device. Exemplary artificial restorations include, withoutlimitation, dental restorations, dentures, dental prosthesis,craniofacial implants, artificial joints, and artificial bones.Exemplary medical devices include, but are not limited to, catheters,medical diagnosis instruments such as endoscopes, contact lenses,spectacles, hearing aids, and mouth guards.

The spinel ferrite nanoparticles may be applied onto a desired area ofthe surface as needed. In certain embodiments, the method disclosedherein involves applying the spinel ferrite nanoparticles onto thesurface 1 to 10 times daily, preferably 2 to 7 times daily, preferably 3to 5 times daily. In some embodiments, the interval of time between eachapplication of the spinel ferrite nanoparticles may be about 1-5minutes, 1-30 minutes, 30 minutes to 60 minutes, 1 hour, 1-2 hours, 2-6hours, 2-12 hours, 12-24 hours. 1-2 days, 2 days, 3 days, 4 days, 5days, 6 days, 7 days, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 15 weeks, 20 weeks, 26weeks, 52 weeks, 11-15 weeks, 15-20 weeks, 20-30 weeks, 30-40 weeks,40-50 weeks, 1 month, 2 months, 3 months, 4 months, 5 months, 6 months,7 months, 8 months, 9 months, 10 months, 11 months, 12 months, 1 year, 2years, or any period of time in between.

In one embodiment, the aforementioned method in any of its embodimentsmay prevent or reduce growth of gram-negative bacteria including, butnot limited to, Escherichia coli, Escherichia fergusonii, Escherichiahermannii, Escherichia vulneris, Shigella boydii, Shigella dysenteriae,Shigella flexneri, Shigella sonnei, Proteus mirabilis, Proteus penneri,Proteus vulgaris, Cronobacter sakazakii, Pantoea agglomerans, Serratiamarcescens, Citrobacter amalonaticus, Citrobacter braakii, Citrobacterfreundii, Citrobacter koseri, Klebsiella granulomatis, Klebsiellaoxytoca, Klebsiella pneumoniae, Klebsiella varicola, Acinetobacterbaumannii, Acinetobacter calcoaceticus, Acinetobactercolistiniresistens, Acinetobacter defluvii, Acinetobacter haemolyticus,Acinetobacter junii, Acinetobacter lwoffii, Acinetobacter pittii,Acinetobacter schindleri, Acinetobacter soli, Enterobacter aerogenes,Enterobacter tavlorae, Pseudomonas aeruginosa, Neisseria meningitidis,Neisseria gonorrhoeae, Pseudomonas pseudomallei, Treponema pallidum,Mycobacterium tuberculosis, Salmonella spec., alpha-Proteobacteria(particularly Agrobacterium spec.), beta-Proteobacteria (particularlyNitrosomonas spec.). Aquabacterium spec., Gammaproteobacteria,Stenotrophomonas spec. (particularly S. maltophilia), Xanthomonas spec.(X. campestris), Neisseria spec., and Haemophilus spec. Pathogenicstrains of Escherichia coli (E. coli) can cause gastroenteritis, urinarytract infections, neonatal meningitis, hemorrhagic colitis, and Crohn'sdisease. Common signs and symptoms include severe abdominal cramps,diarrhea, hemorrhagic colitis, vomiting, and sometimes fever. In atleast one embodiment, the method disclosed herein in any of itsembodiments may prevent or reduce growth of E. coli.

The method disclosed herein in any of its embodiments may be effectiveon pathogenic gram-positive bacteria including, but not limited to,Staphylococcus aureus, MRSA, Staphylococcus auricularis, Staphylococcuscapitis, Staphylococcus caprae, Staphylococcus cohnii, Staphylococcusdelphini, Staphylococcus epidermis, Staphylococcus felis, Staphylococcusgallinarum, Staphylococcus haemolyticus, Staphylococcus hominis,Staphylococcus hyicus, Staphylococcus lugdunensis, Staphylococcuspettenkoferi, Staphylococcus pseudintermedius, Staphylococcus rostri,Staphylococcus saccharolvticus, Staphylococcus saprophyticus,Staphylococcus schleiferi, Staphylococcus vitulinus, Staphylococcuswarneri, Staphylococcus xylosus, Streptococcus pyogenes, Corynebacteriumspp. (particularly C. tenuis, C. diphtheriae, and C. minutissimum),Micrococcus spp. (particularly M. sedentarius), Bacillus anthracis,Streptococcus spec. (particularly S. gordonii, and S. mutans),Actinomyces spec. (particularly A. naeslundii), and Actinobacteria(particularly Brachybacterium spec.).

In certain embodiments, preventing or reducing growth of a microorganismon the surface may be evaluated by measuring microbial counts of thesurface before and/or at least 30 minutes, preferably at least 1 hour,more preferably at least 2 hours after applying the surface with thespinel ferrite nanoparticles via the method described herein in any ofits embodiments. For example, the number of viable microorganisms iscounted using a slide count method and/or a direct culture method (platecount).

The “slide count” method utilizes a microscope slide in a chamber thatis especially designed to enable cell counting. A total number of cellsin a sample can be determined by looking at the sample under amicroscope and counting the number manually. A number of viable cellscan also be determined using the slide count method if a viability dyeis added to the sample. Exemplary viability dyes include, but are notlimited to. Trypan Blue, Calcein-AM, Erythrosine B, propidium iodide,and 7-aminoactinomycin D.

“Colony-forming unit (CFU)” refers to a unit used to estimate the numberof viable bacteria or fungal cells in a sample. The purpose of directculture method (plate count) is to estimate the number of cells presentbased on their ability to give rise to colonies under specificconditions of nutrient medium, temperature and time. Theoretically, oneviable cell can give rise to a colony through replication. A samplesolution of microbes at an unknown concentration is often seriallydiluted in order to obtain at least one plate with a countable number ofCFUs. Counting colonies is performed manually using a pen and aclick-counter, or automatically using an automated system and a softwaretool for counting CFUs.

Preventing or reducing growth of a microorganism on a surface may beunderstood to indicate a reduction of the number of microorganism cellson the surface after applying the spinel ferrite nanoparticles onto thesurface. In some embodiments, the number of microorganisms on thesurface characterized by a microbial count is reduced by at least 10%,preferably at least 20%, more preferably at least 30%, more preferablyat least 40%, more preferably at least 45%, more preferably at least50%, more preferably at least 55%, more preferably at least 60%, morepreferably at least 65%, more preferably at least 70%, more preferablyat least 80%, more preferably at least 90%, or more preferably at least95%, with respect to use of an untreated control surface. Ideally, thegrowth of microorganisms on the surface may be completely or almostcompletely prevented.

As defined herein, the minimum inhibitory concentration (MIC) of anantimicrobial composition for a given microorganism is the lowestconcentration of the composition required to inhibit the growth of themicroorganism. The minimum bactericidal concentration (MBC) is thelowest concentration of the antimicrobial composition required to killthe microorganism. In general, an antimicrobial composition is regardedas bactericidal if the MBC is no more than four times the MIC.

In one embodiment, the antimicrobial efficacy of the spinel ferritenanoparticles comprising at least one chromium-substituted copperferrite selected from the group consisting of CuCr_(0.4)Fe_(1.6)O₄,CuCr_(0.6)Fe_(1.4)O₄, CuCr_(0.8)Fe_(1.2)O₄, and CuCrFeO₄ is demonstratedby test data showing the minimum inhibitory concentrations (MIC) andminimum bactericidal concentrations (MBC) against microorganisms (e.g.E. coli) cultured in vitro under standard conditions (see Examples8-13).

In a preferred embodiment, the spinel ferrite nanoparticles comprisingat least one chromium-substituted copper ferrite selected from the groupconsisting of CuCr_(0.4)Fe_(1.6)O₄, CuCr_(0.6)Fe_(1.4)O₄,CuCr_(0.8)Fe_(1.2)O₄, and CuCrFeO₄ have a MIC against E. coli that is atleast 50% smaller than the MIC against E. coli of substantially similarspinel ferrite nanoparticles comprising a copper ferrite lackingchromium substitution (e.g. CuFe₂O₄), preferably at least 60% smaller,preferably at least 70% smaller, preferably at least 75% smaller,preferably at least 80% smaller, preferably at least 85% smaller,preferably at least 87.5% smaller, preferably at least 90% smaller,preferably at least 95% smaller than the MIC against E. coli ofsubstantially similar spinel ferrite nanoparticles comprising a copperferrite lacking chromium substitution (see Table 3). In a relatedembodiment, the spinel ferrite nanoparticles comprising at least onechromium-substituted copper ferrite selected from the group consistingof CuCr_(0.4)Fe_(1.6)O₄, CuCr_(0.6)Fe_(1.4)O₄, CuCr_(0.8)Fe_(1.2)O₄, andCuCrFeO₄ in any of their embodiments have a MBC against E. coli that isat least 50% smaller than the MBC against E. coli of substantiallysimilar spinel ferrite nanoparticles comprising a copper ferrite lackingchromium substitution (e.g. CuFe₂O₄), preferably at least 60% smaller,preferably at least 70% smaller, preferably at least 75% smaller,preferably at least 80% smaller, preferably at least 85% smaller thanthe MBC against E. coli of substantially similar spinel ferritenanoparticles comprising a copper ferrite lacking chromium substitution(see Table 3).

The examples below are intended to further illustrate protocols forpreparing, characterizing spinel ferrite nanoparticles, and usesthereof, and are not intended to limit the scope of the claims.

Example 1

Synthesis of Chromium-Substituted Copper Ferrite Nanoparticles

Chromium-substituted copper ferrite nanoparticles with a chemicalcomposition of CuCr_(x)Fe_(2-x)O₄, where x=0.0, 0.2, 0.4, 0.6, 0.8 and1.0, were prepared by co-precipitation method using copper(II) nitratepentahydrate (Cu(NO₃)₂·5H₂O), chromium(III) chloride hexahydrate(CrCl₂·6H₂O), iron nitrate (Fe(NO₃)₃), and sodium hydroxide (NaOH) asprecursor reagents. The precursor materials were dissolved in 100 mLdistilled water by applying a constant stirring for 40 min at atemperature of 80° C. The pH of the reaction mixture was maintained at11 using NaOH. The precipitation process was continued for 3 h at 130°C. and then the reaction mixture was dried by heating at 70° C. for 6 h.Finally, all the synthesized products were annealed by heating at 400°C. for 4 h.

Example 2

Characterization Methods

The phase and crystallographic analysis of synthesized ferritenanoparticles were carried out using CuKα radiation (λ=1.514 Å) with anXRD equipment built by Rigaku D/Max-IIIC Diffractometer (Japan). Thecrystalline size of each composition was calculated using Sherrer'sformula [Ansari M A, Khan H M, Khan A A, Sultan A, Azam A. Synthesis andcharacterization of the antibacterial potential of ZnO nanoparticlesagainst extended-spectrum β-lactamases-producing Escherichia coli andKlebsiella pneumoniae isolated from a tertiary care hospital of NorthIndia. Applied microbiology and biotechnology. 2012; 94(2):467-77]. Theexperimental lattice parameter (a) was calculated using the formuladescribed by Klug et al. [Klug H P, L. E. Alexander, X-ray DiffractionProcedures For polycrystalline and Amorphous Materials, John Wiley &Sons, Inc., New York, 1954, L. Lutterotti, P. Scardi, J. Appl.Crystallogr. 1990 23:246-252].

X-ray density, apparent density, porosity, and effect of substitution ofCr³⁺ hopping length between the two sub-lattice sites have beenestimated using the formula described by Batoo et al. [Batoo K M, G.Kumar, Y. Yang, Y. Al-Douri, M. Singh, R. B. Jotania, A. Imran, J.Alloys and Compd. 2017, 726:179-186]. The spectral analysis of thesamples was measured by FT-IR (Bruker) spectrophotometer. Opticalproperties of the samples (diffuse reflectance, DR) were conducted usinga UV-Vis spectrophotometer (Evolution 300 PC Thermo Scientific) equippedwith Praying Mantis Diffuse Reflectance accessory. The optical energyband gap (E_(g)) of synthesized nanoparticles was assessed by applyingthe Kubelka-Munk (K-M) model [Baykal A, S. Esir, A. Demir, S. Goner,Magnetic and optical properties of Cu_(1-x)Zn_(x)Fe₂O₄ nanoparticlesdispersed in a silica matrix by a sol-gel auto-combustion method,Ceramics International. 2015, 41:231-239; and Barathiraja C, A.Manikandan. A. M. Uduman Mohideen, S. Jayasree, S. A. Antony,Magnetically recyclable spinel Mn_(x)Ni_(1-x)Fe₂O₄ (x=0.0-0.5)nano-photocatalysts: Structural, morphological and opto-magneticproperties, Journal of Superconductivity and Novel Magnetism. 2016,29:477-486, each incorporated herein by reference in their entirety]. DR% measurements can be used to determine the absorption coefficient (α)using the formula given below:

${{F(R)} \equiv \alpha} = \frac{\left( {1 - R} \right)^{2}}{2R}$where F(R) is the Kubelka-Munk function, (α) is absorption coefficient,and R is reflectance.

The morphology of the ferrite nanoparticles was examined by SEM (JEOLJSM-6490). The textural properties and nitrogen adsorption/desorptionisotherms of as-prepared nanoparticles were measured using liquidnitrogen (77 K) by Micromeritics ASAP 2020 automatic analyzer. Beforeperforming the analysis, each sample was heated at 150° C. for 3 h toeliminate unwanted gas. BET analysis was used to determine the surfacearea while the pore size and volume distribution data were obtained byapplying BJH method.

Example 3

X-ray Diffraction Analysis of Synthesized CuCr_(x)Fe_(2-x)O₄ (0.0≤x≤1.0)Nanoparticles (NPs)

X-ray diffraction patterns of synthesized CuCr_(x)Fe_(2-x)O₄ (0.0≤x≤1.0)NPs were analyzed using powder-X software (FIGS. 1A-F). The most intensereflection peak (311) was observed at 34.8°, which is a characteristicpeak of spinel phase. Further, the peaks (220), (311), (222), (400),(511), (440), and (531) indicate the presence of a cubic spinel phasewith the space group Fd₃m (Samavati A, Ismail A F. Antibacterialproperties of copper-substituted cobalt ferrite nanoparticlessynthesized by co-precipitation method. Particuology. 2017; 30:158-63;and Ashour A H, El-Batal A I, Abdel Maksoud M I A, El-Sayyad G S, LabibcS, Abdeltwab E, El-Okr M M. Antimicrobial activity of metal-substitutedcobalt ferrite nanoparticles synthesized by sol-gel technique.Particuology. 2018; volume 40, pages 141-151, each incorporated hereinby reference in their entirety]. The estimated crystallite sizesobtained from the Scherer plots were ˜43.3, ˜25.6, ˜34.5, ˜27.5, 22.9,and ˜20.2 nm for x=0.00, 0.2, 0.4, 0.6, 0.8, and 1, respectively (Table1). It has been found that the crystallite size decreased linearly withincreasing substitution of the dopant ion (Cr), obeying Vegard's law.The results obtained are in good agreement with those of Samavati andIsmail (2017) where they found that the crystallite size decreased withincreasing Cu content in copper-substituted cobalt ferrite nanoparticlessynthesized by co-precipitation method. It was also observed that thelattice parameter increased linearly as the concentration of Cr³⁺ ionsincreased (Table 1). Both X-ray and bulk densities were found toincrease slightly with the substitution of the Cr³⁺ ions. The slightincrease in porosity could be reasoned on the basis that the density ofthe Cr³⁺ ion (7.19 g/cm³) is less than the density of the replacingelement Fe³⁺ ion (7.874 g/cm³), which caused decreases in X-ray as wellas apparent density and an increase in porosity (Table 1). It was notedthat the hopping length was affected by substitution of Cr³⁺ as thehopping length increased at both A- and B-Sites slightly upon thesubstitution. The above observation may be resulted from the fact thatCr³⁺ ions have an occupational preference for octahedral B-site, howeverwhen the concentration of Cr³⁺ ions increases to a certain limit, theions can also occupy tetrahedral A-site, as reported in the literature[Haralkar S. J., R. H. Kadam, S. S. More, Sagar E. Shirsath, S. Patil,D. R. Mane, Phys. B, Cond. Matt. 2012 407:4338-4346; and More S. S, R.H. Kadam, A. B. Kadam, A. R. Shite, D. R. Mane, K. M. Jadhav, J. Alloy.Compds. 2010, 502:477-479, each incorporated herein by reference intheir entirety].

TABLE 1 Cr content and structural parameters of CuCr_(x)Fe_(2-x)O₄ (0.0≤ x ≤ 1.0) NPs Crystallite Lattice X-ray Sample size Parameter densityTh. Density Porosity Hopping Length x (nm) (a) (Å) (ρ_(x)) (g/cm³) (ρ)(g/cm³) (%) A-site B-site 0 43.33 8.485 14.85 11.56 22.15 3.6525 2.98220.2 25.65 8.4856 14.83 11.42 22.99 3.6527 2.9824 0.4 34.58 8.4856 14.5511.28 22.47 3.6743 3.0001 0.6 27.54 8.4865 14.53 11.02 24.15 3.67473.0004 0.8 22.98 8.4869 14.5 10.95 24.68 3.6749 3.0005 1 20.21 8.487314.5 10.82 25.37 3.6751 3.0007

Example 4

FT-IR Analysis of Synthesized CuCr_(x)Fe_(2-x)O₄ (0.0≤x≤1.0) NPs

FT-IR spectral analyses of synthesized spinel CuCr_(x)Fe_(2-x)O₄(0.0≤x≤1.0) NPs recorded in the range of 400-4000 cm⁻¹ were shown inFIG. 2 . The spinel ferrite nanoparticles often exhibit two FTIR-activebands, designated as w₁ (higher absorption band) and w₂ (lowerabsorption band) in the range of 450-900 cm⁻¹ [Padmapriya G, ManikandanA, Krishnasamy V, Jaganathan S K, Antony S A. Spinel Ni_(x)Zn_(1-x)Fe₂O₄(0.0≤x≤1.0) nano-photocatalysts: synthesis, characterization andphotocatalytic degradation of methylene blue dye. Journal of MolecularStructure. 2016; 1119:39-47, incorporated herein by reference in itsentirety]. In the present disclosure, FT-IR spectra of all the samplesexhibit two intense peaks at ˜500 cm⁻¹ (lower peak) and ˜850 cm⁻¹(higher peak) corresponding to metal oxygen (Cu—O, Cr—O, and Fe—O) bonds[Padmapriya G, Manikandan A, Krishnasamy V, Jaganathan S K, Antony S A.Spinel Ni_(x)Zn_(1-x)Fe₂O₄ (0.0≤x≤1.0) nano-photocatalysts: synthesis,characterization and photocatalytic degradation of methylene blue dye.Journal of Molecular Structure. 2016; 1119:39-47; Silambarasu A,Manikandan A, Balakrishnan K. Room-Temperature Superparamagnetism andEnhanced Photocatalytic Activity of Magnetically Reusable Spinel ZnFe₂O₄Nanocatalysts. Journal of Superconductivity and Novel Magnetism. 2017;30(9):2631-40; and Suguna S, Shankar S, Jaganathan S K, Manikandan A.Novel synthesis of spinel Mn_(x)Co_(1-x)Al₂O₄ (x=0.0 to 1.0)nanocatalysts: effect of Mn²⁺ doping on structural, morphological, andopto-magnetic properties. Journal of Superconductivity and NovelMagnetism. 2017; 30(3):691-9, each incorporated herein by reference intheir entirety]. The significant Fe—O vibrational mode clearlydemonstrated the existence of strong Cr doping in CuFe₂O₄ spinel lattice[Teresita V M, Manikandan A, Josephine B A, Sujatha S, Antony S A.Electromagnetic Properties and Humidity-Sensing Studies of MagneticallyRecoverable LaMg_(x)Fe_(1-x)O_(3-δ) Perovskites Nano-photocatalysts bySol-Gel Route. Journal of Superconductivity and Novel Magnetism. 2016,29(6):1691-701; and Josephine B A, Manikandan A, Teresita V M, Antony SA. Fundamental study of LaMg_(x)Cr_(1-x)O_(3-δ) perovskitesnano-photocatalysts: Sol-gel synthesis, characterization and humiditysensing. Korean Journal of Chemical Engineering. 2016; 33(5):1590-8,each incorporated herein by reference in their entirety].

Example 5

UV-Visible Diffuse Reflectance Spectroscopy Analysis

The optical properties of spinel CuCr_(x)Fe_(2-x)O₄ (0.0≤x≤1.0) NPs wereinvestigated by UV-vis percent diffuse reflectance (DR %) spectroscopy.The recorded spectra had a sweep range of 200-800 nm (FIG. 3A). Allsamples absorbed at least 72% or more of the light from 200 to 600 nmrange. Above 600 nm, DR remarkably increased at different magnitudesdepending on doped Cr ratio. Cr ratio dependent band gaps (E_(g)) wereshown in FIG. 3B. The E_(g) values were found in the range of 1.20 eV to1.80 eV for CuCr_(x)Fe_(2-x)O₄ (x=0.0, 0.2, 0.4, 0.6, 0.8, 1.0) NPs(FIG. 3B). The estimated band gap value increased from 1.52 eV ofCuFe₂O₄ to a maximum of 1.80 eV for x=0.6, and a sharp decrease to aminimum of 1.20 eV for x=1.0 (FIG. 3B), which might be due to thesmaller particle size.

Example 6

Analysis of Surface Area and Pore Size Distribution: BET and BJHMeasurements

N₂ adsorption/desorption isotherms (BET) and BJH desorption pore sizedistribution plots of the synthesized spinel Cr substituted ferritenanoparticles were shown in FIG. 4 and FIG. 5 , respectively. Over thelast three decades, BET analysis has been used to measure the specificsurface area and texture of a wide range of porous materials [Sing K.The use of nitrogen adsorption for the characterization of porousmaterials. Colloids and Surfaces A: Physicochemical and EngineeringAspects 187-188 (2001) 3-9]. The isotherm curves of all samples showed atypical Langmuir IV type, with hysteresis according to IUPACclassification, disclosing the existence of a mesoporous structure[Ashour A H, El-Batal A I, Abdel Maksoud M I A, El-Sayyad G S, Labibc S,Abdeltwab E, El-Okr M M. Antimicrobial activity of metal-substitutedcobalt ferrite nanoparticles synthesized by sol-gel technique.Particuology. 2018; volume 40, pages 141-151]. The BET surface area ofCr substituted ferrite nanoparticles decreased as Cr content increased,which might be attributed to the increase in grain size (Table 2). FIG.5 shows the BJH desorption average pore diameter (d_(p)) of Crsubstituted ferrite nanoparticles and it was observed that the porediameter increased going from CuFe₂O₄ to CuCrFeO₂ (FIG. 5 ). Varioustextural and structural properties of synthesized spinel ferritenanoparticles were summarized in Table 2.

TABLE 2 Surface properties of the spinel ferrite nanoparticles S_(BET)S_(micro) ^(a)) S_(meso) ^(b)) V_(t) ^(c)) V_(micro) ^(a)) d_(p) ^(d))APS^(e)) Sample x (m²g⁻¹) (m²g⁻¹) (m²g⁻¹) (cm³g⁻¹) (cm³g⁻¹) (nm) (nm)HF^(f)) CuFe₂O₄ 0 19.6 1.5 18.1 0.02 0.02 8.7 65.0 0.92CuCr_(0.2)Fe_(1.8)O₄ 0.2 18.8 1.7 17.1 0.019 0.01 12.4 73.5 0.47CuCr_(0.4)Fe_(1.6)O₄ 0.4 14 1.1 12.9 0.03 0.01 10.4 47.6 0.31CuCr_(0.6)Fe_(1.4)O₄ 0.6 13.5 1.0 12.5 0.02 0.01 11 69.9 0.46CuCr_(0.8)Fe_(1.2)O₄ 0.8 11.1 0.8 10.3 0.02 0.03 10.7 54.2 1.4 CuCrFeO₄1 10.5 0.6 9.9 0.03 0.03 14.5 28.0 0.94 ^(a))Micropore area/volumecalculated using the t-plot; ^(b))External surface area calculated usingthe t-plot; ^(c))Total pore volume adsorbed at p/p⁰ = 0.95; ^(d))BJHAdsorption average pore width (4 V/A); ^(e))Average particle size (APS):^(f))Hierarchical factor (HF), HF = (V_(micro)/V_(total)) *(S_(meso)/S_(BET))

Example 7

Scanning Electron Microscopy Analysis

The morphology of nanoparticles of CuCr_(x)Fe_(2-x)O₄ ferrites wasexamined by SEM. FIG. 6 shows the SEM images of the synthesizedCr-substituted copper ferrite nanoparticles (A) CuFe₂O₄, (B)CuCr_(0.2)Fe_(1.8)O₄, (C) CuCr_(0.4)Fe_(1.6)O₄, (D)CuCr_(0.6)Fe_(1.4)O₄, (E) CuCr_(0.8)Fe_(1.2)O₄, and (F) CuCrFeO₄). TheCuFe₂O₄(FIG. 6A), CuCr_(0.2)Fe_(1.8)O₄(FIG. 6B), and CuCrFeO₄ (FIG. 6F)nanoparticles exhibit spherical morphology. While FIGS. 6C-E presentparticles with angular, spinel, and irregular shapes. The averageparticle sizes of CuFe₂O₄, (B) CuCr_(0.2)Fe_(1.8)O₄, (C)CuCr_(0.4)Fe_(1.6)O₄, (D) CuCr_(0.6)Fe_(1.4)O₄, (E) CuCr_(0.8)Fe_(1.2)O₄and (F) CuCrFeO₄ were ˜65.0, ˜73.5, ˜47.6, ˜69.9, 54.2 and ˜28.0 nm,respectively. The SEM micrographs showed that the samples were porous innature, with porosities observed between 22-25% as confirmed by XRD.

Example 8

Characterizations of Antibacterial Activity of Spinel Nanoparticles

E. coli were chosen as model Gram-negative bacteria to investigate theantibacterial properties of CuCr_(x)Fe_(2-x)O₄ (0.0≤x≤1.0) NPs. E. colicultures were grown overnight in nutrient broth medium in a shakingincubator (200 rpm) at 37° C. The bacterial culture was then washed 2-3times with phosphate buffered saline and the E. coli suspensions werediluted with sterile 0.9% NaCl solution to reach a final concentrationof approximately 10⁷ CFU/mL.

Example 9

Minimal Inhibitory Concentration (MIC)

The antibacterial activity of CuCr_(x)Fe_(2-x)O₄ (0.0≤x≤1.0) NPs wasassessed using the standard agar dilution method as previously described[Ansari M A, Khan H M, Alzohairy M A, Jalal M, Ali S G, Pal R, MusarratJ. Green synthesis of Al₂O₃ nanoparticles and their bactericidalpotential against clinical isolates of multi-drug resistant Pseudomonasaeruginosa. World Journal of Microbiology and Biotechnology. 2015,31(1):153-64]. The MIC was determined on MHA (Mueller Hinton Agar)plates using serial dilutions of CuCr_(x)Fe_(2-x)O₄ (0.0≤x≤1.0) NPs atconcentration ranges from 32 mg/mL to 0.5 mg/mL. The MIC is determinedas the lowest concentration of NPs at which no visible growth of thebacteria was observed [Ansari M A, Khan H M, Alzohairy M A, Jalal M, AliS G, Pal R, Musarrat J. Green synthesis of Al₂O₃ nanoparticles and theirbactericidal potential against clinical isolates of multi-drug resistantPseudomonas aeruginosa. World Journal of Microbiology and Biotechnology.2015, 31(1):153-64].

Example 10

Minimal Bactericidal Concentration (MBC)

The concentrations of CuCr_(x)Fe_(2-x)O₄ (0.0≤x≤1.0) NPs showedeffective inhibition of bacteria growth were selected for MBCexamination [Jalal M, Ansari M A, Shukla A K, Ali S G, Khan H M, Pal R,Alam J, Cameotra S S. Green synthesis and antifungal activity of Al₂O₃NPs against fluconazole-resistant Candida spp isolated from a tertiarycare hospital. RSC Advances. 2016, 6(109):107577-90]. Briefly, 100 μL0.9% normal saline were added onto the MIC plates, which was transferredto another freshly prepared MHA plate and then incubated at 37° C. for24 h [Jalal et al. 2016]. The lowest concentration of CuCr_(x)Fe_(2-x)O₄(0.0≤x≤1.0) NPs at which no growth of bacterial cells found or less thanthree CFUs present was recorded as MBC [Jalal et al. 2016].

Example 11

Effect of Spinel Ferrite Nanoparticles on the Morphology of E. coli: SEMAnalysis

The antibacterial effect of CuCr_(x)Fe_(2-x)O₄ (0.0≤x≤1.0) NPs on themorphology of E. coli cells was investigated by scanning electronmicroscope using previously reported methods [Ansari M A, Khan H M,Alzohairy M A, Jalal M, Ali S G, Pal R, Musarrat J. Green synthesis ofAl₂O₃ nanoparticles and their bactericidal potential against clinicalisolates of multi-drug resistant Pseudomonas aeruginosa. World Journalof Microbiology and Biotechnology. 2015, 31(1):153-64; and Jalal M,Ansari M A, Shukla A K, Ali S G, Khan H M, Pal R, Alam J, Cameotra S S.Green synthesis and antifungal activity of Al₂O₃ NPs againstfluconazole-resistant Candida spp isolated from a tertiary carehospital. RSC Advances. 2016, 6(109):107577-90]. Briefly, ˜10⁶ CFU/mL ofE. coli cells were treated with 2 mg/mL of CuCr_(x)Fe_(2-x)O₄(0.0≤x≤1.0) NPs at 37° C. for 12 h. After centrifugation at 12000 rpmfor 10 min, pellets were obtained and then washed at least three timeswith PBS, initially fixed with 2.5% glutaraldehyde, and further fixedwith 1% osmium tetroxide. After washing, the samples were dehydrated bya series of ethanol solvent [Ansari M A, Khan H M, Alzohairy M A, JalalM, Ali S G, Pal R, Musarrat J. Green synthesis of Al₂O₃ nanoparticlesand their bactericidal potential against clinical isolates of multi-drugresistant Pseudomonas aeruginosa. World Journal of Microbiology andBiotechnology. 2015, 31(1):153-64; and Jalal M, Ansari M A, Shukla A K,Ali S G, Khan H M, Pal R, Alam J, Cameotra S S. Green synthesis andantifungal activity of Al₂O₃ NPs against fluconazole-resistant Candidaspp isolated from a tertiary care hospital. RSC Advances. 2016,6(109):107577-90]. The cells were then fixed on the aluminum stubs,dried in a desecrator, and coated with gold to prepare SEM samples.Finally, these samples were examined at an accelerating voltage of 20 kVby SEM.

Example 12

Antibacterial Activity of Chromium-Substituted Copper FerriteNanoparticles

In the present disclosure, antibacterial properties ofchromium-substituted copper ferrite nanoparticles against E. coli (ATCC25922) have been evaluated by determining MICs and MBCs using agardilution methods. The MICs and MBCs values of spinel CuCr_(x)Fe_(2-x)O₄(x=0.0, 0.2, 0.4, 0.6, 0.8, 1.0) nanoparticles were summarized in Table3. The lowest MIC and MBC values recorded were 2.5 and 5 mg/mL forCuCr_(0.4)Fe_(1.6)O₄. CuCr_(0.8)Fe_(1.2)O₄ and CuCrFeO₄ nanoparticles,whereas CuFe₂O₄ and CuCr_(0.2)Fe_(1.8)O₄ nanoparticles showed larger MICand MBC values, i.e., >16 and >32 mg/mL. In a recent study, Ashour etal. [Ashour A H, El-Batal A I, Abdel Maksoud M I A, El-Sayyad G S,Labibc S, Abdeltwab E, El-Okr M M. Antimicrobial activity ofmetal-substituted cobalt ferrite nanoparticles synthesized by sol-geltechnique. Particuology. 2018; volume 40, pages 141-151, incorporatedherein by reference in its entirety] reported that metal-substitutedcobalt ferrite nanoparticles such as copper cobalt ferrite, zinc cobaltferrite and manganese cobalt ferrite inhibited growth of E. coli and S.aureus at a concentration of 5000 ppm. It has also been found that asthe content of dopant (Cr) increased from 0.0 to 1.0, the antibacterialactivity of ferrite nanoparticles also increased (Table 3). In aprevious study, it was reported that the bactericidal activity ofcopper-substituted cobalt ferrite nanoparticles enhanced when thecontent of Cu increased [Samavati A, Ismail A F. Antibacterialproperties of copper-substituted cobalt ferrite nanoparticlessynthesized by co-precipitation method. Particuology. 2017; 30:158-63,incorporated herein by reference in its entirety]. Further, it wasobserved that the CuCr_(x)Fe_(2-x)O₄ nanoparticles inhibited bacterialgrowth in a size dependent manner i.e., smaller size CuCr_(x)Fe_(2-x)O₄(20.2 nm; x=1.0) nanoparticles exhibited stronger antibacterial activityand inhibited bacterial growth at lower concentrations i.e., 2.5 mg/mL,whereas larger size CuCr_(x)Fe₂O₄ nanoparticles (43.3 nm; x=0.0)inhibited bacterial growth at higher concentrations i.e., >16 mg/mL(Table 3). Results on size dependent antimicrobial activity of chromiumsubstituted copper ferrite nanoparticles were in good agreement ofprevious study by Žalnėravičius et al. [Žalnėravičius R, Paškevičius A,Kurtinaitiene M, Jagminas A. Size-dependent antimicrobial properties ofthe cobalt ferrite nanoparticles. Journal of Nanoparticle Research.2016, 18(10):300, incorporated herein by reference in its entirety],where they also reported size dependent antimicrobial activity of cobaltferrite nanoparticles. Size dependent antimicrobial activity has alsobeen reported for metal oxide nanoparticles such as ZnO [Raghupathi K R,Koodali R T, Manna A C. Size-dependent bacterial growth inhibition andmechanism of antibacterial activity of zinc oxide nanoparticles.Langmuir. 2011, 27(7):4020-8, incorporated herein by reference in itsentirety] and CuO [She B, Wan X, Tang J, Deng Y, Zhou X, Xiao C. Size-and Morphology-Dependent Antibacterial Properties of Cuprous OxideNanoparticle and Their Synergistic Antibacterial Effect. Science ofAdvanced Materials. 2016, 8(5):1074-8, incorporated herein by referencein its entirety]. The content of chromium ions (factor of x=0.2) in theouter part of copper ferrite NP shell and effect on number of smallersized particles was disclosed [Žalnėravičius R, Paškevičius A,Kurtinaitiene M, Jagminas A. Size-dependent antimicrobial properties ofthe cobalt ferrite nanoparticles. Journal of Nanoparticle Research.2016, 18(10):300, incorporated herein by reference in its entirety].

TABLE 3 MIC and MBC (mg/mL) values of chromium- substituted copperferrite nanoparticles (CuCr_(x)Fe₂-_(x)O₄ where x = 0.0, 0.2, 0.4, 0.6,0.8, and 1.0) against E. coli Chromium-substituted Crystallite MIC MBCMBC/ copper size (mg/ (mg/ MIC ferrite nanoparticles (nm) mL) mL) ratioCuFe₂O₄ 43.33 >16 >32 >2 CuCr0.2Fe_(1.8)O₄ 25.65 >16 >32 >2CuCr0.4Fe_(1.6)O₄ 34.58  2.5  5  2 CuCr0.6Fe_(1.4)O₄ 27.54  4  8  2CuCr0.8Fe_(1.2)O₄ 22.98  2.5  5  2 CuCrFeO₄ 20.21  2.5  5  2

Example 13

Effects of CuCr_(x)Fe_(2-x)O₄ Nanoparticles on the Morphology of E.coli: SEM Analysis

The morphological and structural changes in E. coli cells caused bychromium-substituted copper ferrite nanoparticles were furtherinvestigated by SEM. The untreated (i.e., control) E. coli cells weretypically rod-shaped and regular with a smooth cell surface. Cell walland cell membrane of untreated E. coli cells were normal and intact(FIG. 7A). However, E. coli cells treated with CuCr_(x)Fe_(2-x)O₄nanoparticles were not intact i.e., abnormal in shape with irregularfragments appeared at the cell surface (FIGS. 7B-G). E. coli cellstreated with the presently disclosed nanoparticles were severely damagedas pits, indentation, deformation, and distortion of cell wall andmembrane were observed, which indicated that significant loss ofmembrane integrity might lead to cell death (FIGS. 7B-G). There is verylittle information available in the literature regarding theantimicrobial properties and mode of action of metal-substituted ferritenanoparticles against bacteria [Sanpo N, Wen C, Berndt C C, Wang J.Antibacterial properties of spinel ferrite nanoparticles. Microbialpathogens and strategies for combating them: science, technology andeducation. Spain: Formatex Research Centre. 2013:239-50; and Samavati A,Ismail A F. Antibacterial properties of copper-substituted cobaltferrite nanoparticles synthesized by co-precipitation method.Particuology. 2017; 30:158-63, each incorporated herein by reference intheir entirety]. Previous reports on other nanoparticles againstbacteria disclosed that morphological and structural alteration in thebacterial cell membrane due to nanoparticles attachment and penetrationmight be a possible mode of action [Jalal M, Ansari M A, Shukla A K, AliS G, Khan H M, Pal R, Alam J. Cameotra S S. Green synthesis andantifungal activity of Al₂O₃ NPs against fluconazole-resistant Candidaspp isolated from a tertiary care hospital. RSC Advances. 2016,6(109):107577-90; Sondi I, Salopek-Sondi B. Silver nanoparticles asantimicrobial agent: a case study on E. coli as a model forGram-negative bacteria. Journal of colloid and interface science. 2004,275(1):177-82; Jiang W, Mashayekhi H, Xing B (2009) Bacterial toxicitycomparison between nano- and micro-scale oxide particles. Environ Pollut157:1619-1625; and Leung Y H, Xu X, Ma A P, Liu F, Ng A M, Shen Z,Gethings L A, Guo M Y, Djurišić A B, Lee P K, Lee H K. Toxicity of ZnOand TiO₂ to Escherichia coli cells. Scientific reports. 2016, 6:35243,each incorporated herein by reference in their entirety]. Thus, theattachments of NPs to bacterial cell surface might play an importantrole in achieving good bactericidal activity. From SEM images (FIGS.7B-G), it was clear that CuCr_(x)Fe_(2-x)O₄ nanoparticles were able toadhere to bacterial cell surfaces and damage the cell membrane due tointeraction between nanoparticles and cell membrane [Jalal M, Ansari MA, Shukla A K, Ali S G, Khan H M, Pal R, Alam J, Cameotra S S. Greensynthesis and antifungal activity of Al₂O₃ NPs againstfluconazole-resistant Candida spp isolated from a tertiary carehospital. RSC Advances. 2016, 6(109):107577-90; and Leung Y H, Xu X, MaA P, Liu F, Ng A M, Shen Z, Gethings L A, Guo M Y, Djurišić A B, Lee PK, Lee H K. Toxicity of ZnO and TiO₂ to Escherichia coli cells.Scientific reports. 2016, 6:35243, each incorporated herein by referencein their entirety]. Another possible mechanism reported in previousstudies was the generation of reactive oxygen species (ROS) bynanoparticles. In the current disclosure, decomposition ofCuCr_(x)Fe_(2-x)O₄ nanoparticles may generate ROS that electrostaticallyinteract with cell walls and cause damage to bacteria cells [Schwartz VB, Thétiot F, Ritz S, Pütz S, Choritz L, Lappas A, Förch R, LandfesterK, Jonas U. Antibacterial surface coatings from zinc oxide nanoparticlesembedded in poly (n-isopropylacrylamide) hydrogel surface layers.Advanced Functional Materials. 2012, 22(11):2376-86, incorporated hereinby reference in its entirety]. The results were in good agreement withthe previous study by Sanpo et al. [Sanpo N, Wen C, Berndt C C, Wang J.Antibacterial properties of spinel ferrite nanoparticles. Microbialpathogens and strategies for combating them: science, technology andeducation. Spain: Formatex Research Centre. 2013:239-50, incorporatedherein by reference in its entirety], which reported that generation ofROS from spinel metal substituted cobalt ferrite nanoparticles had morepotential to enter the cell wall and inhibit the growth of E. coli andS. aureus. In previous studies. ROS generated from the surface of NPsmay have interacted with the cell membrane and damaged the membrane dueto increased cell permeability and leakage of the intracellularmaterials [Leung Y H, Xu X, Ma A P, Liu F, Ng A M, Shen Z, Gethings L A,Guo M Y, Djurišić A B, Lee P K, Lee H K. Toxicity of ZnO and TiO₂ toEscherichia coli cells. Scientific reports. 2016, 6:35243; Schwartz V B,Thetiot F, Ritz S, Pütz S, Choritz L, Lappas A, Förch R, Landfester K,Jonas U. Antibacterial surface coatings from zinc oxide nanoparticlesembedded in poly (n-isopropylacrylamide) hydrogel surface layers.Advanced Functional Materials. 2012, 22(11):2376-86; and Pandey B K,Shahi A K, Srivastava N, Kumar G, Gopal R (2015) Synthesis andcytogenetic effect of magnetic nanoparticles. Adv Mater Lett 6:954-960,each incorporated herein by reference in their entirety]. Tran et al.[Tran N, Mir A, Mallik D, Sinha A, Nayar A, Webster T J (2010)Bactericidal effect of iron oxide nanoparticles on Staphylococcusaureus. Int J Nanomed 5:277-283, incorporated herein by reference in itsentirety] also reported that Fe₂O₃ NPs could penetrate bacterial cellsand generate ROS. Further, as the concentration of dopant (Cr) increasedin CuCr_(x)Fe_(2-x)O₄ nanoparticles, smaller crystalline sizednanoparticles were obtained with a lager surface-to-volume ratio. Ahigher surface area of CuCr_(x)Fe_(2-x)O₄ nanoparticles might enhancetheir antimicrobial activity and killing rates [Samavati A, Ismail A F.Antibacterial properties of copper-substituted cobalt ferritenanoparticles synthesized by co-precipitation method. Particuology.2017; 30:158-63; and Stoimenov P K, Klinger R L, Marchin G L, Klabunde KJ. Metal oxide nanoparticles as bactericidal agents. Langmuir. 2002,18(17):6679-86, each incorporated herein by reference in theirentirety].

Example 14

Chromium-substituted copper ferrite nanoparticles were prepared byco-precipitation method. All the chromium-substituted nanoparticles werespinel in structure. Substitution of chromium ions has influenced thecrystal and microstructure of copper ferrite nanoparticles and enhancedtheir antibacterial property. Further, as the content of Cr ionsincreases from 0.0 to 1.0, the crystallite size of copper ferritenanoparticles decreases from 43.3 to 20.2 nm. Size dependentantibacterial activity of chromium-substituted copper ferritenanoparticles has been investigated for the first time. CuCrFeO₄nanoparticles with smaller size (20.2 nm) exhibit MIC of 2.5 mg/mL,whereas larger size CuFe₂O₄ (43.3 nm) nanoparticles show MIC of >16mg/mL. Because of their excellent antibacterial activity, Cr-substitutedcopper ferrite nanoparticles may be used in coating of medical devicesto prevent microbial biofilm growth, magnetic drug delivery systems, aswell as ointments, cosmetics, creams and lotions for topicalapplication.

The invention claimed is:
 1. Spinel ferrite nanoparticles, comprising atleast one chromium-substituted copper ferrite selected from the groupconsisting of CuCr_(0.4)Fe_(1.6)O₄, CuCr_(0.6)Fe_(1.4)O₄,CuCr_(0.8)Fe_(1.2)O₄, and CuCrFeO₄.
 2. The spinel ferrite nanoparticlesof claim 1, comprising a chromium-substituted copper ferrite having theformula CuCr_(0.4)Fe_(1.6)O₄.
 3. The spinel ferrite nanoparticles ofclaim 1, comprising a chromium-substituted copper ferrite having theformula CuCr_(0.6)Fe_(1.4)O₄.
 4. The spinel ferrite nanoparticles ofclaim 1, comprising a chromium-substituted copper ferrite having theformula CuCr_(0.8)Fe_(1.2)O₄.
 5. The spinel ferrite nanoparticles ofclaim 1, consisting of the chromium-substituted copper ferrite.
 6. Thespinel ferrite nanoparticles of claim 1, having a crystallite size offrom 43.3 to 20.2 nm.
 7. The spinel ferrite nanoparticles of claim 1,having spherical morphology and an average particle size of from 73.5 to47.6 nm.
 8. The spinel ferrite nanoparticles of claim 1, wherein thespinel ferrite nanoparticles have an average particle size in a range of20-90 nm.
 9. The spinel ferrite nanoparticles of claim 1, wherein thespinel ferrite nanoparticles are porous.
 10. The spinel ferritenanoparticles of claim 1, wherein the spinel ferrite nanoparticles havea BET surface area in a range of 8-30 m²/g.
 11. The spinel ferritenanoparticles of claim 1, wherein the spinel ferrite nanoparticles havean optical band gap energy value of 1.0-2.0 eV.